Antibodies to hsdek49 polypeptides

ABSTRACT

The present invention relates to human secreted polypeptides, and isolated nucleic acid molecules encoding said polypeptides, useful for diagnosing and treating immune disorders and diseases. Antibodies that bind these polypeptides are also encompassed by the present invention. Also encompassed by the invention are vectors, host cells, and recombinant and synthetic methods for producing said polynucleotides, polypeptides, and/or antibodies. The invention further encompasses screening methods for identifying agonists and antagonists of polynucleotides and polypeptides of the invention. The present invention further encompasses methods and compositions for inhibiting or enhancing the production and function of the polypeptides of the present invention.

This application is a divisional of U.S. application Ser. No. 11/346,470, filed Feb. 3, 2006, which is a continuation-in-part of U.S. application Ser. No. 10/472,532, filed Sep. 20, 2003 and accorded a National Stage filing date of Jun. 23, 2004, now abandoned, which is the 35 U.S.C. §371 National Stage of PCT/US02/08278, filed Mar. 19, 2002, which in turn claims benefit of U.S. patent application Ser. No. 10/100,683 (now U.S. Pat. No. 7,368,531, issued May 6, 2008). U.S. application Ser. No. 11/346,470 is also a continuation-in-part of U.S. patent application Ser. No. 10/100,683, filed Mar. 19, 2002, which in turn claims benefit of the following:

Application:: Continuity Type:: Parent Application:: Parent Filing Date:: 10/100,683 Non-provisional of 60/277,340 Mar. 21, 2001 10/100,683 Non-provisional of 60/306,171 Jul. 19, 2001 10/100,683 Non-provisional of 60/331,287 Nov. 13, 2001 10/100,683 Continuation-in-part of 09/981,876 Oct. 19, 2001 09/981,876 Divisional of 09/621,011 Jul. 20, 2000 09/621,011 Continuation of 09/148,545 Sep. 04, 1998 09/148,545 Continuation-in-part of PCT/US98/04482 Mar. 06, 1998 10/100,683 Continuation-in-part of 09/621,011 Jul. 20, 2000 09/621,011 Continuation of 09/148,545 Sep. 04, 1998 09/148,545 Continuation-in-part of PCT/US98/04482 Mar. 06, 1998 10/100,683 Continuation-in-part of 09/148,545 Sep. 04, 1998 09/148,545 Continuation-in-part of PCT/US98/04482 Mar. 06, 1998 10/100,683 Continuation-in-part of PCT/US98/04482 Mar. 06, 1998 PCT/US98/04482 Non-provisional of 60/040,162 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/040,333 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/038,621 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/040,161 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/040,626 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/040,334 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/040,336 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/040,163 Mar. 07, 1997 PCT/US98/04482 Non-provisional of 60/047,615 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,600 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,597 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,502 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,633 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,583 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,617 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,618 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,503 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,592 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,581 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,584 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,500 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,587 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,492 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,598 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,613 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,582 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,596 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,612 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,632 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,601 May 23, 1997 PCT/US98/04482 Non-provisional of 60/043,580 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,568 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,314 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,569 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,311 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,671 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,674 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,669 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,312 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,313 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,672 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,315 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/048,974 Jun. 06, 1997 PCT/US98/04482 Non-provisional of 60/056,886 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,877 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,889 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,893 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,630 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,878 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,662 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,872 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,882 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,637 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,903 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,888 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,879 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,880 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,894 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,911 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,636 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,874 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,910 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,864 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,631 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,845 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,892 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/047,595 May 23, 1997 PCT/US98/04482 Non-provisional of 60/057,761 Sep. 05, 1997 PCT/US98/04482 Non-provisional of 60/047,599 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,588 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,585 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,586 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,590 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,594 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,589 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,593 May 23, 1997 PCT/US98/04482 Non-provisional of 60/047,614 May 23, 1997 PCT/US98/04482 Non-provisional of 60/043,578 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/043,576 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/047,501 May 23, 1997 PCT/US98/04482 Non-provisional of 60/043,670 Apr. 11, 1997 PCT/US98/04482 Non-provisional of 60/056,632 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,664 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,876 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,881 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,909 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,875 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,862 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,887 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/056,908 Aug. 22, 1997 PCT/US98/04482 Non-provisional of 60/048,964 Jun. 06, 1997 PCT/US98/04482 Non-provisional of 60/057,650 Sep. 05, 1997 PCT/US98/04482 Non-provisional of 60/056,884 Aug. 22, 1997 10/100,683 Continuation-in-part of 09/882,171 Jun. 18, 2001 09/882,171 Non-provisional of 60/190,068 Mar. 17, 2000 09/882,171 Continuation of 09/809,391 Mar. 16, 2001 09/809,391 Continuation-in-part of 09/149,476 Sep. 08, 1998 09/149,476 Continuation-in-part of PCT/US98/04493 Mar. 06, 1998 10/100,683 Continuation-in-part of 09/809,391 Mar. 16, 2001 09/809,391 Non-provisional of 60/190,068 Mar. 17, 2000 09/809,391 Continuation-in-part of 09/149,476 Sep. 08, 1998 09/149,476 Continuation-in-part of PCT/US98/04493 Mar. 06, 1998 10/100,683 Continuation-in-part of 09/149,476 Sep. 08, 1998 09/149,476 Continuation-in-part of PCT/US98/04493 Mar. 06, 1998 10/100,683 Continuation-in-part of PCT/US98/04493 Mar. 06, 1998 PCT/US98/04493 Non-provisional of 60/040,161 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/040,162 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/040,333 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/038,621 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/040,626 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/040,334 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/040,336 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/040,163 Mar. 07, 1997 PCT/US98/04493 Non-provisional of 60/047,600 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,615 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,597 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,502 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,633 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,583 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,617 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,618 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,503 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,592 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,581 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,584 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,500 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,587 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,492 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,598 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,613 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,582 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,596 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,612 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,632 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,601 May 23, 1997 PCT/US98/04493 Non-provisional of 60/043,580 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,568 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,314 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,569 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,311 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,671 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,674 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,669 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,312 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,313 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,672 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,315 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/048,974 Jun. 06, 1997 PCT/US98/04493 Non-provisional of 60/056,886 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,877 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,889 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,893 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,630 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,878 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,662 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,872 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,882 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,637 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,903 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,888 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,879 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,880 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,894 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,911 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,636 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,874 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,910 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,864 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,631 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,845 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,892 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/057,761 Sep. 05, 1997 PCT/US98/04493 Non-provisional of 60/047,595 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,599 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,588 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,585 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,586 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,590 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,594 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,589 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,593 May 23, 1997 PCT/US98/04493 Non-provisional of 60/047,614 May 23, 1997 PCT/US98/04493 Non-provisional of 60/043,578 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/043,576 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/047,501 May 23, 1997 PCT/US98/04493 Non-provisional of 60/043,670 Apr. 11, 1997 PCT/US98/04493 Non-provisional of 60/056,632 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,664 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,876 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,881 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,909 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,875 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,862 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,887 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/056,908 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/048,964 Jun. 06, 1997 PCT/US98/04493 Non-provisional of 60/057,650 Sep. 05, 1997 PCT/US98/04493 Non-provisional of 60/056,884 Aug. 22, 1997 PCT/US98/04493 Non-provisional of 60/057,669 Sep. 05, 1997 PCT/US98/04493 Non-provisional of 60/049,610 Jun. 13, 1997 PCT/US98/04493 Non-provisional of 60/061,060 Oct. 02, 1997 PCT/US98/04493 Non-provisional of 60/051,926 Jul. 08, 1997 PCT/US98/04493 Non-provisional of 60/052,874 Jul. 16, 1997 PCT/US98/04493 Non-provisional of 60/058,785 Sep. 12, 1997 PCT/US98/04493 Non-provisional of 60/055,724 Aug. 18, 1997 10/100,683 Continuation-in-part of 10/058,993 Jan. 30, 2002 10/058,993 Non-provisional of 60/265,583 Feb. 02, 2001 10/058,993 Continuation-in-part of 09/852,659 May 11, 2001 09/852,659 Continuation-in-part of 09/152,060 Sep. 11, 1998 09/152,060 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 10/058,993 Continuation-in-part of 09/853,161 May 11, 2001 09/853,161 Continuation-in-part of 09/152,060 Sep. 11, 1998 09/152,060 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 10/058,993 Continuation-in-part of 09/852,797 May 11, 2001 09/852,797 Continuation-in-part of 09/152,060 Sep. 11, 1998 09/152,060 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 10/100,683 Continuation-in-part of 09/852,659 May 11, 2001 09/852,659 Non-provisional of 60/265,583 Feb. 02, 2001 09/852,659 Continuation-in-part of 09/152,060 Sep. 11, 1998 09/152,060 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 10/100,683 Continuation-in-part of 09/853,161 May 11, 2001 09/853,161 Non-provisional of 60/265,583 Feb. 02, 2001 09/853,161 Continuation-in-part of 09/152,060 Sep. 11, 1998 09/152,060 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 10/100,683 Continuation-in-part of 09/852,797 May 11, 2001 09/852,797 Non-provisional of 60/265,583 Feb. 02, 2001 09/852,797 Continuation-in-part of 09/152,060 Sep. 11, 1998 09/152,060 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 10/100,683 Continuation-in-part of 09/152,060 Sep. 11, 1998 09/152,060 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 10/100,683 Continuation-in-part of PCT/US98/04858 Mar. 12, 1998 PCT/US98/04858 Non-provisional of 60/040,762 Mar. 14, 1997 PCT/US98/04858 Non-provisional of 60/040,710 Mar. 14, 1997 PCT/US98/04858 Non-provisional of 60/050,934 May 30, 1997 PCT/US98/04858 Non-provisional of 60/048,100 May 30, 1997 PCT/US98/04858 Non-provisional of 60/048,357 May 30, 1997 PCT/US98/04858 Non-provisional of 60/048,189 May 30, 1997 PCT/US98/04858 Non-provisional of 60/057,765 Sep. 05, 1997 PCT/US98/04858 Non-provisional of 60/048,970 Jun. 06, 1997 PCT/US98/04858 Non-provisional of 60/068,368 Dec. 19, 1997 10/100,683 Continuation-in-part of 10/059,395 Jan. 31, 2002 10/059,395 Divisional of 09/966,262 Oct. 01, 2001 09/966,262 Continuation of 09/154,707 Sep. 17, 1998 09/154,707 Continuation-in-part of PCT/US98/05311 Mar. 19, 1998 10/100,683 Continuation-in-part of 09/984,245 Oct. 29, 2001 09/984,245 Divisional of 09/154,707 Sep. 17, 1998 09/154,707 Continuation-in-part of PCT/US98/05311 Mar. 19, 1998 10/100,683 Continuation-in-part of 09/983,966 Oct. 26, 2001 09/983,966 Divisional of 09/154,707 Sep. 17, 1998 09/154,707 Continuation-in-part of PCT/US98/05311 Mar. 19, 1998 10/100,683 Continuation-in-part of 09/966,262 Oct. 01, 2001 09/966,262 Continuation of 09/154,707 Sep. 17, 1998 09/154,707 Continuation-in-part of PCT/US98/05311 Mar. 19, 1998 10/100,683 Continuation-in-part of 09/154,707 Sep. 17, 1998 09/154,707 Continuation-in-part of PCT/US98/05311 Mar. 19, 1998 10/100,683 Continuation-in-part of PCT/US98/05311 Mar. 03, 1998 PCT/US98/05311 Non-provisional of 60/041,277 Mar. 21, 1997 PCT/US98/05311 Non-provisional of 60/042,344 Mar. 21, 1997 PCT/US98/05311 Non-provisional of 60/041,276 Mar. 21, 1997 PCT/US98/05311 Non-provisional of 60/041,281 Mar. 21, 1997 PCT/US98/05311 Non-provisional of 60/048,094 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,350 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,188 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,135 May 30, 1997 PCT/US98/05311 Non-provisional of 60/050,937 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,187 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,099 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,352 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,186 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,069 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,095 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,131 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,096 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,355 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,160 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,351 May 30, 1997 PCT/US98/05311 Non-provisional of 60/048,154 May 30, 1997 PCT/US98/05311 Non-provisional of 60/054,804 Aug. 05, 1997 PCT/US98/05311 Non-provisional of 60/056,370 Aug. 19, 1997 PCT/US98/05311 Non-provisional of 60/060,862 Oct. 02, 1997 10/100,683 Continuation-in-part of 09/814,122 Mar. 22, 2001 09/814,122 Continuation of 09/577,145 May 24, 2000 09/577,145 Continuation of 09/166,780 Oct. 06, 1998 09/166,780 Continuation-in-part of PCT/US98/06801 Apr. 07, 1998 10/100,683 Continuation-in-part of PCT/US98/06801 Apr. 07, 1998 PCT/US98/06801 Non-provisional of 60/042,726 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,727 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,728 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,754 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,825 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/048,068 May 30, 1997 PCT/US98/06801 Non-provisional of 60/048,070 May 30, 1997 PCT/US98/06801 Non-provisional of 60/048,184 May 30, 1997 10/100,683 Continuation-in-part of PCT/US98/06801 Apr. 07, 1997 PCT/US98/06801 Non-provisional of 60/042,726 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,727 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,728 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,754 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/042,825 Apr. 08, 1997 PCT/US98/06801 Non-provisional of 60/048,068 May 30, 1997 PCT/US98/06801 Non-provisional of 60/048,070 May 30, 1997 PCT/US98/06801 Non-provisional of 60/048,184 May 30, 1997 10/100,683 Continuation-in-part of PCT/US98/10868 May 28, 1998 PCT/US98/10868 Non-provisional of 60/044,039 May 30, 1997 PCT/US98/10868 Non-provisional of 60/048,093 May 30, 1997 PCT/US98/10868 Non-provisional of 60/048,190 May 30, 1997 PCT/US98/10868 Non-provisional of 60/050,935 May 30, 1997 PCT/US98/10868 Non-provisional of 60/048,101 May 30, 1997 PCT/US98/10868 Non-provisional of 60/048,356 May 30, 1997 PCT/US98/10868 Non-provisional of 60/056,250 Aug. 29, 1997 PCT/US98/10868 Non-provisional of 60/056,296 Aug. 29, 1997 PCT/US98/10868 Non-provisional of 60/056,293 Aug. 29, 1997 10/100,683 Continuation-in-part of PCT/US98/11422 Jun. 04, 1998 PCT/US98/11422 Non-provisional of 60/048,885 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/049,375 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,881 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,880 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,896 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/049,020 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,876 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,895 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,884 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,894 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,971 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,964 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,882 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,899 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,893 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,900 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,901 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,892 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,915 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/049,019 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,970 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,972 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,916 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/049,373 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,875 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/049,374 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,917 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,949 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,974 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,883 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,897 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,898 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,962 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,963 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,877 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/048,878 Jun. 06, 1997 PCT/US98/11422 Non-provisional of 60/057,645 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,642 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,668 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,635 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,627 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,667 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,666 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,764 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,643 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,769 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,763 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,650 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,584 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,647 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,661 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,662 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,646 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,654 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,651 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,644 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,765 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,762 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,775 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,648 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,774 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,649 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,770 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,771 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,761 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,760 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,776 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,778 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,629 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,628 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,777 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/057,634 Sep. 05, 1997 PCT/US98/11422 Non-provisional of 60/070,923 Dec. 18, 1997 10/100,683 Continuation-in-part of PCT/US01/05614 Feb. 21, 2001 PCT/US01/05614 Non-provisional of 60/184,836 Feb. 24, 2000 PCT/US01/05614 Non-provisional of 60/193,170 Mar. 29, 2000 10/100,683 Continuation-in-part of PCT/US98/12125 Jun. 11, 1998 PCT/US98/12125 Non-provisional of 60/049,547 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,548 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,549 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,550 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,566 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,606 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,607 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,608 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,609 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,610 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/049,611 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/050,901 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/052,989 Jun. 13, 1997 PCT/US98/12125 Non-provisional of 60/051,919 Jul. 08, 1997 PCT/US98/12125 Non-provisional of 60/055,984 Aug. 18, 1997 PCT/US98/12125 Non-provisional of 60/058,665 Sep. 12, 1997 PCT/US98/12125 Non-provisional of 60/058,668 Sep. 12, 1997 PCT/US98/12125 Non-provisional of 60/058,669 Sep. 12, 1997 PCT/US98/12125 Non-provisional of 60/058,750 Sep. 12, 1997 PCT/US98/12125 Non-provisional of 60/058,971 Sep. 12, 1997 PCT/US98/12125 Non-provisional of 60/058,972 Sep. 12, 1997 PCT/US98/12125 Non-provisional of 60/058,975 Sep. 12, 1997 PCT/US98/12125 Non-provisional of 60/060,834 Oct. 02, 1997 PCT/US98/12125 Non-provisional of 60/060,841 Oct. 02, 1997 PCT/US98/12125 Non-provisional of 60/060,844 Oct. 02, 1997 PCT/US98/12125 Non-provisional of 60/060,865 Oct. 02, 1997 PCT/US98/12125 Non-provisional of 60/061,059 Oct. 02, 1997 PCT/US98/12125 Non-provisional of 60/061,060 Oct. 02, 1997 10/100,683 Continuation-in-part of 09/627,081 Jul. 27, 2000 09/627,081 Continuation of 09/213,365 Dec. 17, 1998 09/213,365 Continuation-in-part of PCT/US98/13608 Jun. 30, 1998 10/100,683 Continuation-in-part of PCT/US98/13608 Jun. 30, 1998 PCT/US98/13608 Non-provisional of 60/051,480 Jul. 01, 1997 PCT/US98/13608 Non-provisional of 60/051,381 Jul. 01, 1997 PCT/US98/13608 Non-provisional of 60/058,663 Sep. 12, 1997 PCT/US98/13608 Non-provisional of 60/058,598 Sep. 12, 1997 10/100,683 Continuation-in-part of 09/984,490 Oct. 30, 2001 09/984,490 Divisional of 09/227,357 Jan. 08, 1999 09/227,357 Continuation-in-part of PCT/US98/13684 Jul. 07, 1998 10/100,683 Continuation-in-part of 09/983,802 Oct. 25, 2001 09/983,802 Continuation of 09/227,357 Oct. 10, 2001 09/227,357 Continuation-in-part of PCT/US98/13684 Jul. 07, 1998 10/100,683 Continuation-in-part of 09/973,278 Oct. 10, 2001 09/973,278 Non-provisional of 60/239,899 Oct. 13, 2000 09/973,278 Continuation-in-part of 09/227,357 Jan. 08, 1999 09/227,357 Continuation-in-part of PCT/US98/13684 Jul. 07, 1998 10/100,683 Continuation-in-part of PCT/US98/13684 Jul. 07, 1998 PCT/US98/13684 Non-provisional of 60/051,926 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/052,793 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,925 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,929 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/052,803 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/052,732 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,931 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,932 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,916 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,930 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,918 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,920 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/052,733 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/052,795 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,919 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/051,928 Jul. 08, 1997 PCT/US98/13684 Non-provisional of 60/055,722 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,723 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,948 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,949 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,953 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,950 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,947 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,964 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/056,360 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,684 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,984 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/055,954 Aug. 18, 1997 PCT/US98/13684 Non-provisional of 60/058,785 Sep. 12, 1997 PCT/US98/13684 Non-provisional of 60/058,664 Sep. 12, 1997 PCT/US98/13684 Non-provisional of 60/058,660 Sep. 12, 1997 PCT/US98/13684 Non-provisional of 60/058,661 Sep. 12, 1997 10/100,683 Continuation-in-part of 09/776,724 Feb. 06, 2001 09/776,724 Non-provisional of 60/180,909 Feb. 08, 2000 09/776,724 Continuation-in-part of 09/669,688 Sep. 26, 2000 09/669,688 Continuation of 09/229,982 Jan. 14, 1999 09/229,982 Continuation-in-part of PCT/US98/14613 Jul. 15, 1998 10/100,683 Continuation-in-part of 09/669,688 Sep. 26, 2000 09/669,688 Continuation of 09/229,982 Jan. 14, 1999 09/229,982 Continuation-in-part of PCT/US98/14613 Jul. 15, 1998 10/100,683 Continuation-in-part of 09/229,982 Jan. 14, 1999 09/229,982 Continuation-in-part of PCT/US98/14613 Jul. 15, 1998 10/100,683 Continuation-in-part of PCT/US98/14613 Jul. 15, 1998 PCT/US98/14613 Non-provisional of 60/052,661 Jul. 16, 1997 PCT/US98/14613 Non-provisional of 60/052,872 Jul. 16, 1997 PCT/US98/14613 Non-provisional of 60/052,871 Jul. 16, 1997 PCT/US98/14613 Non-provisional of 60/052,874 Jul. 16, 1997 PCT/US98/14613 Non-provisional of 60/052,873 Jul. 16, 1997 PCT/US98/14613 Non-provisional of 60/052,870 Jul. 16, 1997 PCT/US98/14613 Non-provisional of 60/052,875 Jul. 16, 1997 PCT/US98/14613 Non-provisional of 60/053,440 Jul. 22, 1997 PCT/US98/14613 Non-provisional of 60/053,441 Jul. 22, 1997 PCT/US98/14613 Non-provisional of 60/053,442 Jul. 22, 1997 PCT/US98/14613 Non-provisional of 60/056,359 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,725 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,985 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,952 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,989 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/056,361 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,726 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,724 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,946 Aug. 18, 1997 PCT/US98/14613 Non-provisional of 60/055,683 Aug. 18, 1997 10/100,683 Non-provisional of 60/295,558 Jun. 05, 2001 10/100,683 Continuation-in-part of 09/820,649 Mar. 30, 2001 09/820,649 Continuation of 09/666,984 Sep. 21, 2000 09/666,984 Continuation of 09/236,557 Jan. 26, 1999 09/236,557 Continuation-in-part of PCT/US98/15949 Jul. 29, 1998 10/100,683 Continuation-in-part of PCT/US98/15949 Jul. 29, 1998 PCT/US98/15949 Non-provisional of 60/054,212 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,209 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,234 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,218 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,214 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,236 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,215 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,211 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,217 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/054,213 Jul. 30, 1997 PCT/US98/15949 Non-provisional of 60/055,968 Aug. 18, 1997 PCT/US98/15949 Non-provisional of 60/055,969 Aug. 18, 1997 PCT/US98/15949 Non-provisional of 60/055,972 Aug. 18, 1997 PCT/US98/15949 Non-provisional of 60/056,561 Aug. 19, 1997 PCT/US98/15949 Non-provisional of 60/056,534 Aug. 19, 1997 PCT/US98/15949 Non-provisional of 60/056,729 Aug. 19, 1997 PCT/US98/15949 Non-provisional of 60/056,543 Aug. 19, 1997 PCT/US98/15949 Non-provisional of 60/056,727 Aug. 19, 1997 PCT/US98/15949 Non-provisional of 60/056,554 Aug. 19, 1997 PCT/US98/15949 Non-provisional of 60/056,730 Aug. 19, 1997 10/100,683 Continuation-in-part of 09/969,730 Oct. 04, 2001 09/969,730 Continuation-in-part of 09/774,639 Feb. 01, 2001 09/774,639 Continuation of 09/244,112 Feb. 04, 1999 09/244,112 Continuation-in-part of PCT/US98/16235 Aug. 04, 1998 10/100,683 Continuation-in-part of 09/774,639 Feb. 01, 2001 09/774,639 Continuation of 09/244,112 Feb. 04, 1999 09/244,112 Continuation-in-part of PCT/US98/16235 Aug. 04, 1998 10/100,683 Continuation-in-part of 09/969,730 Oct. 04, 2001 09/969,730 Non-provisional of 60/238,291 Oct. 06, 2000 10/100,683 Continuation-in-part of PCT/US98/16235 Aug. 04, 1998 PCT/US98/16235 Non-provisional of 60/055,386 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/054,807 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/055,312 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/055,309 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/054,798 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/055,310 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/054,806 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/054,809 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/054,804 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/054,803 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/054,808 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/055,311 Aug. 05, 1997 PCT/US98/16235 Non-provisional of 60/055,986 Aug. 18, 1997 PCT/US98/16235 Non-provisional of 60/055,970 Aug. 18, 1997 PCT/US98/16235 Non-provisional of 60/056,563 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,557 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,731 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,365 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,367 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,370 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,364 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,366 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,732 Aug. 19, 1997 PCT/US98/16235 Non-provisional of 60/056,371 Aug. 19, 1997 10/100,683 Continuation-in-part of 09/716,128 Nov. 17, 2000 09/716,128 Continuation of 09/251,329 Feb. 17, 1999 09/251,329 Continuation-in-part of PCT/US98/17044 Aug. 18, 1998 10/100,683 Continuation-in-part of PCT/US98/17044 Aug. 18, 1998 PCT/US98/17044 Non-provisional of 60/056,555 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,556 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,535 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,629 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,369 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,628 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,728 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,368 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/056,726 Aug. 19, 1997 PCT/US98/17044 Non-provisional of 60/089,510 Jun. 16, 1998 PCT/US98/17044 Non-provisional of 60/092,956 Jul. 15, 1998 10/100,683 Continuation-in-part of 09/729,835 Dec. 06, 2000 09/729,835 Divisional of 09/257,179 Feb. 25, 1999 09/257,179 Continuation-in-part of PCT/US98/17709 Aug. 27, 1998 10/100,683 Continuation-in-part of 09/257,179 Feb. 25, 1999 09/257,179 Continuation-in-part of PCT/US98/17709 Aug. 27, 1998 10/100,683 Continuation-in-part of PCT/US98/17709 Aug. 27, 1998 PCT/US98/17709 Non-provisional of 60/056,270 Aug. 29, 1997 PCT/US98/17709 Non-provisional of 60/056,271 Aug. 29, 1997 PCT/US98/17709 Non-provisional of 60/056,247 Aug. 29, 1997 PCT/US98/17709 Non-provisional of 60/056,073 Aug. 29, 1997 10/100,683 Continuation-in-part of 10/047,021 Jan. 17, 2002 10/047,021 Continuation-in-part of 09/722,329 Nov. 28, 2000 09/722,329 Continuation of 09/262,109 Mar. 04, 1999 09/262,109 Continuation-in-part of PCT/US98/18360 Sep. 03, 1998 10/100,683 Continuation-in-part of 09/722,329 Nov. 28, 2000 09/722,329 Continuation of 09/262,109 Mar. 04, 1999 09/262,109 Continuation-in-part of PCT/US98/18360 Sep. 03, 1998 10/100,683 Continuation-in-part of PCT/US02/01109 Jan. 17, 2002 PCT/US02/01109 Non-provisional of 60/262,066 Jan. 18, 2001 10/100,683 Continuation-in-part of PCT/US98/18360 Sep. 03, 1998 PCT/US98/18360 Non-provisional of 60/057,626 Sep. 05, 1997 PCT/US98/18360 Non-provisional of 60/057,663 Sep. 05, 1997 PCT/US98/18360 Non-provisional of 60/057,669 Sep. 05, 1997 PCT/US98/18360 Non-provisional of 60/058,667 Sep. 12, 1997 PCT/US98/18360 Non-provisional of 60/058,974 Sep. 12, 1997 PCT/US98/18360 Non-provisional of 60/058,973 Sep. 12, 1997 PCT/US98/18360 Non-provisional of 60/058,666 Sep. 12, 1997 PCT/US98/18360 Non-provisional of 60/090,112 Jun. 22, 1998 10/100,683 Continuation-in-part of 09/281,976 Mar. 31, 1999 09/281,976 Continuation-in-part of PCT/US98/20775 Oct. 01, 1998 10/100,683 Continuation-in-part of PCT/US98/20775 Oct. 01, 1998 PCT/US98/20775 Non-provisional of 60/060,837 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,862 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,839 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,866 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,843 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,836 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,838 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,874 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,833 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,884 Oct. 02, 1997 PCT/US98/20775 Non-provisional of 60/060,880 Oct. 02, 1997 10/100,683 Continuation-in-part of 09/984,429 Oct. 30, 2001 09/984,429 Non-provisional of 60/244,591 Nov. 01, 2000 09/984,429 Continuation-in-part of 09/288,143 Apr. 08, 1999 09/288,143 Continuation-in-part of PCT/US98/21142 Oct. 08, 1998 10/100,683 Non-provisional of 60/244,591 Nov. 01, 2000 10/100,683 Continuation-in-part of 09/288,143 Apr. 08, 1999 09/288,143 Continuation-in-part of PCT/US98/21142 Oct. 08, 1998 10/100,683 Continuation-in-part of PCT/US98/21142 Oct. 08, 1998 PCT/US98/21142 Non-provisional of 60/061,463 Oct. 09, 1997 PCT/US98/21142 Non-provisional of 60/061,529 Oct. 09, 1997 PCT/US98/21142 Non-provisional of 60/071,498 Oct. 09, 1997 PCT/US98/21142 Non-provisional of 60/061,527 Oct. 09, 1997 PCT/US98/21142 Non-provisional of 60/061,536 Oct. 09, 1997 PCT/US98/21142 Non-provisional of 60/061,532 Oct. 09, 1997 10/100,683 Continuation-in-part of 09/296,622 Apr. 23, 1999 09/296,622 Continuation-in-part of PCT/US98/22376 Oct. 23, 1998 10/100,683 Continuation-in-part of PCT/US98/22376 Oct. 23, 1998 PCT/US98/22376 Non-provisional of 60/063,099 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,088 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,100 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,387 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,148 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,386 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/062,784 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,091 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,090 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,089 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,092 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,111 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,101 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,109 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,110 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,098 Oct. 24, 1997 PCT/US98/22376 Non-provisional of 60/063,097 Oct. 24, 1997 10/100,683 Continuation-in-part of 09/974,879 Oct. 12, 2001 09/974,879 Non-provisional of 60/239,893 Oct. 13, 2000 09/974,879 Continuation-in-part of 09/818,683 Mar. 28, 2001 09/818,683 Continuation of 09/305,736 May 05, 1999 09/305,736 Continuation-in-part of PCT/US98/23435 Nov. 04, 1998 10/100,683 Continuation-in-part of 09/818,683 Mar. 28, 2001 09/818,683 Continuation of 09/305,736 May 05, 1999 09/305,736 Continuation-in-part of PCT/US98/23435 Nov. 04, 1998 10/100,683 Continuation-in-part of 09/305,736 May 05, 1999 09/305,736 Continuation-in-part of PCT/US98/23435 Nov. 04, 1998 10/100,683 Continuation-in-part of PCT/US98/23435 Nov. 04, 1998 PCT/US98/23435 Non-provisional of 60/064,911 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,912 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,983 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,900 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,988 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,987 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,908 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,984 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/064,985 Nov. 07, 1997 PCT/US98/23435 Non-provisional of 60/066,094 Nov. 17, 1997 PCT/US98/23435 Non-provisional of 60/066,100 Nov. 17, 1997 PCT/US98/23435 Non-provisional of 60/066,089 Nov. 17, 1997 PCT/US98/23435 Non-provisional of 60/066,095 Nov. 17, 1997 PCT/US98/23435 Non-provisional of 60/066,090 Nov. 17, 1997 10/100,683 Continuation-in-part of 09/334,595 Jun. 17, 1999 09/334,595 Continuation-in-part of PCT/US98/27059 Dec. 17, 1998 10/100,683 Continuation-in-part of PCT/US98/27059 Dec. 17, 1998 PCT/US98/27059 Non-provisional of 60/070,923 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,007 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,057 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,006 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,369 Dec. 19, 1997 PCT/US98/27059 Non-provisional of 60/068,367 Dec. 19, 1997 PCT/US98/27059 Non-provisional of 60/068,368 Dec. 19, 1997 PCT/US98/27059 Non-provisional of 60/068,169 Dec. 19, 1997 PCT/US98/27059 Non-provisional of 60/068,053 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,064 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,054 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,008 Dec. 18, 1997 PCT/US98/27059 Non-provisional of 60/068,365 Dec. 19, 1997 10/100,683 Continuation-in-part of 09/938,671 Aug. 27, 2001 09/938,671 Continuation of 09/739,907 Dec. 20, 2000 09/739,907 Continuation of 09/348,457 Jul. 07, 1999 09/348,457 Continuation-in-part of PCT/US99/00108 Jan. 06, 1999 10/100,683 Continuation-in-part of 09/739,907 Dec. 20, 2000 09/739,907 Continuation of 09/348,457 Jul. 07, 1999 09/348,457 Continuation-in-part of PCT/US99/00108 Jan. 06, 1999 10/100,683 Continuation-in-part of 09/348,457 Jul. 07, 1999 09/348,457 Continuation-in-part of PCT/US99/00108 Jan. 06, 1999 10/100,683 Continuation-in-part of PCT/US99/00108 Jan. 06, 1999 PCT/US99/00108 Non-provisional of 60/070,704 Jan. 07, 1998 PCT/US99/00108 Non-provisional of 60/070,658 Jan. 07, 1998 PCT/US99/00108 Non-provisional of 60/070,692 Jan. 07, 1998 PCT/US99/00108 Non-provisional of 60/070,657 Jan. 07, 1998 10/100,683 Continuation-in-part of 09/949,925 Sep. 12, 2001 09/949,925 Non-provisional of 60/232,150 Sep. 12, 2000 09/949,925 Continuation-in-part of PCT/US99/01621 Jan. 27, 1999 09/949,925 Continuation-in-part of 09/363,044 Jul. 29, 1999 09/363,044 Continuation-in-part of PCT/US99/01621 Jan. 27, 1999 10/100,683 Continuation-in-part of 09/813,153 Mar. 21, 2001 09/813,153 Continuation of 09/363,044 Jul. 29, 1999 09/363,044 Continuation-in-part of PCT/US99/01621 Jan. 27, 1999 10/100,683 Continuation-in-part of 09/363,044 Jul. 29, 1999 09/363,044 Continuation-in-part of PCT/US99/01621 Jan. 27, 1999 10/100,683 Continuation-in-part of PCT/US99/01621 Jan. 27, 1999 PCT/US99/01621 Non-provisional of 60/073,170 Jan. 30, 1998 PCT/US99/01621 Non-provisional of 60/073,167 Jan. 30, 1998 PCT/US99/01621 Non-provisional of 60/073,165 Jan. 30, 1998 PCT/US99/01621 Non-provisional of 60/073,164 Jan. 30, 1998 PCT/US99/01621 Non-provisional of 60/073,162 Jan. 30, 1998 PCT/US99/01621 Non-provisional of 60/073,161 Jan. 30, 1998 PCT/US99/01621 Non-provisional of 60/073,160 Jan. 30, 1998 PCT/US99/01621 Non-provisional of 60/073,159 Jan. 30, 1998 10/100,683 Continuation-in-part of 10/062,548 Feb. 05, 2002 10/062,548 Continuation of 09/369,247 Aug. 05, 1999 09/369,247 Continuation-in-part of PCT/US99/02293 Feb. 04, 1999 10/100,683 Continuation-in-part of 09/369,247 Aug. 05, 1999 09/369,247 Continuation-in-part of PCT/US99/02293 Feb. 04, 1999 10/100,683 Continuation-in-part of PCT/US99/02293 Feb. 04, 1999 PCT/US99/02293 Non-provisional of 60/074,118 Feb. 09, 1998 PCT/US99/02293 Non-provisional of 60/074,157 Feb. 09, 1998 PCT/US99/02293 Non-provisional of 60/074,037 Feb. 09, 1998 PCT/US99/02293 Non-provisional of 60/074,141 Feb. 09, 1998 PCT/US99/02293 Non-provisional of 60/074,341 Feb. 09, 1998 10/100,683 Continuation-in-part of 09/716,129 Nov. 17, 2000 09/716,129 Continuation-in-part of PCT/US99/03939 Feb. 24, 1999 09/716,129 CON 09/382,572 Aug. 25, 1999 09/382,572 Continuation-in-part of PCT/US99/03939 Feb. 24, 1999 10/100,683 Continuation-in-part of PCT/US99/03939 Feb. 24, 1999 PCT/US99/03939 Non-provisional of 60/076,053 Feb. 26, 1998 PCT/US99/03939 Non-provisional of 60/076,051 Feb. 26, 1998 PCT/US99/03939 Non-provisional of 60/076,054 Feb. 26, 1998 PCT/US99/03939 Non-provisional of 60/076,052 Feb. 26, 1998 PCT/US99/03939 Non-provisional of 60/076,057 Feb. 26, 1998 10/100,683 Continuation-in-part of 09/798,889 Mar. 06, 2001 09/798,889 CON 09/393,022 Sep. 09, 1999 09/393,022 Continuation-in-part of PCT/US99/05721 Mar. 11, 1999 10/100,683 Continuation-in-part of PCT/US99/05721 Mar. 11, 1999 PCT/US99/05721 Non-provisional of 60/077,714 Mar. 12, 1998 PCT/US99/05721 Non-provisional of 60/077,686 Mar. 12, 1998 PCT/US99/05721 Non-provisional of 60/077,687 Mar. 12, 1998 PCT/US99/05721 Non-provisional of 60/077,696 Mar. 12, 1998 10/100,683 Continuation-in-part of 09/397,945 Sep. 17, 1999 09/397,945 Continuation-in-part of PCT/US99/05804 Mar. 18, 1999 10/100,683 Continuation-in-part of PCT/US99/05804 Mar. 18, 1999 PCT/US99/05804 Non-provisional of 60/078,566 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/078,576 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/078,573 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/078,574 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/078,579 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/080,314 Apr. 01, 1998 PCT/US99/05804 Non-provisional of 60/080,312 Apr. 01, 1998 PCT/US99/05804 Non-provisional of 60/078,578 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/078,581 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/078,577 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/078,563 Mar. 19, 1998 PCT/US99/05804 Non-provisional of 60/080,313 Apr. 01, 1998 10/100,683 Continuation-in-part of 09/948,783 Sep. 10, 2001 09/948,783 Non-provisional of 60/231,846 Sep. 11, 2000 09/948,783 Continuation-in-part of 09/892,877 Jun. 28, 2001 09/892,877 Continuation of 09/437,658 Nov. 10, 1999 09/437,658 Continuation-in-part of PCT/US99/09847 May 06, 1999 10/100,683 Continuation-in-part of 09/892,877 Jun. 28, 2001 09/892,877 Continuation of 09/437,658 Nov. 10, 1999 09/437,658 Continuation-in-part of PCT/US99/09847 May 06, 1999 10/100,683 Continuation-in-part of PCT/US99/09847 May 06, 1999 PCT/US99/09847 Non-provisional of 60/085,093 May 12, 1998 PCT/US99/09847 Non-provisional of 60/085,094 May 12, 1998 PCT/US99/09847 Non-provisional of 60/085,105 May 12, 1998 PCT/US99/09847 Non-provisional of 60/085,180 May 12, 1998 PCT/US99/09847 Non-provisional of 60/085,927 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,906 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,920 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,924 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,922 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,923 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,921 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,925 May 18, 1998 PCT/US99/09847 Non-provisional of 60/085,928 May 18, 1998 10/100,683 Continuation-in-part of 10/050,873 Jan. 18, 2002 10/050,873 Non-provisional of 60/263,681 Jan. 24, 2001 10/050,873 Non-provisional of 60/263,230 Jan. 23, 2001 10/050,873 Continuation-in-part of 09/461,325 Dec. 14, 1999 09/461,325 Continuation-in-part of PCT/US99/13418 Jun. 15, 1999 10/100,683 Continuation-in-part of 10/012,542 Dec. 12, 2001 10/012,542 Divisional of 09/461,325 Dec. 14, 1999 09/461,325 Continuation-in-part of PCT/US99/13418 Jun. 15, 1999 10/100,683 Continuation-in-part of 09/461,325 Dec. 14, 1999 09/461,325 Continuation-in-part of PCT/US99/13418 Jun. 15, 1999 10/100,683 Continuation-in-part of PCT/US99/13418 Jun. 15, 1999 PCT/US99/13418 Non-provisional of 60/089,507 Jun. 16, 1998 PCT/US99/13418 Non-provisional of 60/089,508 Jun. 16, 1998 PCT/US99/13418 Non-provisional of 60/089,509 Jun. 16, 1998 PCT/US99/13418 Non-provisional of 60/089,510 Jun. 16, 1998 PCT/US99/13418 Non-provisional of 60/090,112 Jun. 22, 1998 PCT/US99/13418 Non-provisional of 60/090,113 Jun. 22, 1998 10/100,683 Continuation-in-part of 09/984,271 Oct. 29, 2001 09/984,271 Divisional of 09/482,273 Jan. 13, 2000 09/482,273 Continuation-in-part of PCT/US99/15849 Jul. 14, 1999 10/100,683 Continuation-in-part of 09/984,276 Oct. 29, 2001 09/984,276 Divisional of 09/482,273 Jan. 13, 2000 09/482,273 Continuation-in-part of PCT/US99/15849 Jul. 14, 1999 10/100,683 Continuation-in-part of 09/482,273 Jan. 13, 2000 09/482,273 Continuation-in-part of PCT/US99/15849 Jul. 14, 1999 10/100,683 Continuation-in-part of PCT/US99/15849 Jul. 14, 1999 PCT/US99/15849 Non-provisional of 60/092,921 Jul. 15, 1998 PCT/US99/15849 Non-provisional of 60/092,922 Jul. 15, 1998 PCT/US99/15849 Non-provisional of 60/092,956 Jul. 15, 1998 10/100,683 Continuation-in-part of PCT/US01/29871 Sep. 24, 2001 PCT/US01/29871 Non-provisional of 60/234,925 Sep. 25, 2000 PCT/US01/29871 Continuation-in-part of PCT/US01/00911 Jan. 12, 2001 10/100,683 Continuation-in-part of PCT/US01/00911 Jan. 12, 2001 PCT/US01/00911 Continuation-in-part of 09/482,273 Jan. 13, 2000 10/100,683 Non-provisional of 60/350,898 Jan. 25, 2002 10/100,683 Continuation-in-part of 09/489,847 Jan. 24, 2000 09/489,847 Continuation-in-part of PCT/US99/17130 Jul. 29, 1999 10/100,683 Continuation-in-part of PCT/US99/17130 Jul. 29, 1999 PCT/US99/17130 Non-provisional of 60/094,657 Jul. 30, 1998 PCT/US99/17130 Non-provisional of 60/095,486 Aug. 05, 1998 PCT/US99/17130 Non-provisional of 60/096,319 Aug. 12, 1998 PCT/US99/17130 Non-provisional of 60/095,454 Aug. 06, 1998 PCT/US99/17130 Non-provisional of 60/095,455 Aug. 06, 1998 10/100,683 Continuation-in-part of 10/054,988 Jan. 25, 2002 10/054,988 Continuation of 09/904,615 Jul. 16, 2001 09/904,615 Continuation of 09/739,254 Dec. 19, 2000 09/739,254 Continuation of 09/511,554 Feb. 23, 2000 09/511,554 Continuation-in-part of PCT/US99/19330 Aug. 24, 1999 10/100,683 Continuation-in-part of 09/904,615 Jul. 16, 2001 09/904,615 Continuation of 09/739,254 Dec. 19, 2000 09/739,254 Continuation of 09/511,554 Feb. 23, 2000 09/511,554 Continuation-in-part of PCT/US99/19330 Aug. 24, 1999 10/100,683 Continuation-in-part of PCT/US99/19330 Aug. 24, 1999 PCT/US99/19330 Non-provisional of 60/097,917 Aug. 25, 1998 PCT/US99/19330 Non-provisional of 60/098,634 Aug. 31, 1998 10/100,683 Continuation-in-part of 09/820,893 Mar. 30, 2001 09/820,893 Continuation of 09/531,119 Mar. 20, 2000 09/531,119 Continuation-in-part of PCT/US99/22012 Sep. 22, 1999 10/100,683 Continuation-in-part of PCT/US99/22012 Sep. 22, 1999 PCT/US99/22012 Non-provisional of 60/101,546 Sep. 23, 1998 PCT/US99/22012 Non-provisional of 60/102,895 Oct. 02, 1998 10/100,683 Continuation-in-part of 09/948,820 Sep. 10, 2001 09/948,820 Continuation of 09/565,391 May 05, 2000 09/565,391 Continuation-in-part of PCT/US99/26409 Nov. 09, 1999 10/100,683 Continuation-in-part of 09/565,391 May 05, 2000 09/565,391 Continuation-in-part of PCT/US99/26409 Nov. 09, 1999 10/100,683 Continuation-in-part of PCT/US99/26409 Nov. 09, 1999 PCT/US99/26409 Non-provisional of 60/108,207 Nov. 12, 1998 10/100,683 Continuation-in-part of 09/895,298 Jul. 02, 2001 09/895,298 Continuation of 09/591,316 Jun. 09, 2000 09/591,316 Continuation-in-part of PCT/US99/29950 Dec. 16, 1999 10/100,683 Continuation-in-part of PCT/US99/29950 Dec. 16, 1999 PCT/US99/29950 Non-provisional of 60/113,006 Dec. 18, 1998 PCT/US99/29950 Non-provisional of 60/112,809 Dec. 17, 1998 10/100,683 Continuation-in-part of 09/985,153 Nov. 01, 2001 09/985,153 Continuation of 09/618,150 Jul. 17, 2000 09/618,150 Continuation-in-part of PCT/US00/00903 Jan. 18, 2000 10/100,683 Continuation-in-part of PCT/US00/00903 Jan. 18, 2000 PCT/US00/00903 Non-provisional of 60/116,330 Jan. 19, 1999 10/100,683 Continuation-in-part of 09/997,131 Nov. 30, 2001 09/997,131 Continuation of 09/628,508 Jul. 28, 2000 09/628,508 Continuation-in-part of PCT/US00/03062 Feb. 08, 2000 10/100,683 Continuation-in-part of PCT/US00/03062 Feb. 08, 2000 PCT/US00/03062 Non-provisional of 60/119,468 Feb. 10, 1999 10/100,683 Continuation-in-part of 10/050,882 Jan. 18, 2002 10/050,882 Continuation of 09/661,453 Sep. 13, 2000 09/661,453 Continuation-in-part of PCT/US00/06783 Mar. 16, 2000 10/100,683 Continuation-in-part of 09/661,453 Sep. 13, 2000 09/661,453 Continuation-in-part of PCT/US00/06783 Mar. 16, 2000 10/100,683 Continuation-in-part of PCT/US00/06783 Mar. 16, 2000 PCT/US00/06783 Non-provisional of 60/125,055 Mar. 18, 1999 10/100,683 Continuation-in-part of 10/050,704 Jan. 18, 2002 10/050,704 Continuation of 09/684,524 Oct. 10, 2000 09/684,524 Continuation-in-part of PCT/US00/08979 Apr. 06, 2000 10/100,683 Continuation-in-part of 09/684,524 Oct. 10, 2000 09/684,524 Continuation-in-part of PCT/US00/08979 Apr. 06, 2000 10/100,683 Continuation-in-part of PCT/US00/08979 Apr. 06, 2000 PCT/US00/08979 Non-provisional of 60/128,693 Apr. 09, 1999 PCT/US00/08979 Non-provisional of 60/130,991 Apr. 26, 1999 10/100,683 Continuation-in-part of 10/042,141 Jan. 11, 2002 10/042,141 Continuation of 09/726,643 Dec. 01, 2000 09/726,643 Continuation-in-part of PCT/US00/15187 Jun. 02, 2000 10/100,683 Continuation-in-part of 09/726,643 Dec. 01, 2000 09/726,643 Continuation-in-part of PCT/US00/15187 Jun. 02, 2000 10/100,683 Continuation-in-part of PCT/US00/15187 Jun. 02, 2000 PCT/US00/15187 Non-provisional of 60/137,725 Jun. 07, 1999 10/100,683 Continuation-in-part of 09/756,168 Jan. 09, 2001 09/756,168 Continuation-in-part of PCT/US00/19735 Jul. 23, 1999 10/100,683 Continuation-in-part of PCT/US00/19735 Jul. 20, 2000 PCT/US00/19735 Non-provisional of 60/145,220 Jul. 23, 1999 10/100,683 Continuation-in-part of 10/060,255 Feb. 01, 2002 10/060,255 Continuation of 09/781,417 Feb. 13, 2001 09/781,417 Continuation-in-part of PCT/US00/22325 Aug. 16, 2000 10/100,683 Continuation-in-part of 09/781,417 Feb. 13, 2001 09/781,417 Continuation-in-part of PCT/US00/22325 Aug. 16, 2000 10/100,683 Continuation-in-part of PCT/US00/22325 Aug. 16, 2000 PCT/US00/22325 Non-provisional of 60/149,182 Aug. 17, 1999 10/100,683 Continuation-in-part of 09/789,561 Feb. 22, 2001 09/789,561 Continuation-in-part of PCT/US00/24008 Aug. 31, 2000 10/100,683 Continuation-in-part of PCT/US00/24008 Aug. 31, 2000 PCT/US00/24008 Non-provisional of 60/152,315 Sep. 03, 1999 PCT/US00/24008 Non-provisional of 60/152,317 Sep. 03, 1999 10/100,683 Continuation-in-part of 09/800,729 Mar. 08, 2001 09/800,729 Continuation-in-part of PCT/US00/26013 Sep. 22, 2000 10/100,683 Continuation-in-part of PCT/US00/26013 Sep. 22, 2000 PCT/US00/26013 Non-provisional of 60/155,709 Sep. 24, 1999 10/100,683 Continuation-in-part of 09/832,129 Apr. 11, 2001 09/832,129 Continuation-in-part of PCT/US00/28664 Oct. 17, 2000 10/100,683 Continuation-in-part of PCT/US00/28664 Oct. 17, 2000 PCT/US00/28664 Non-provisional of 60/163,085 Nov. 02, 1999 PCT/US00/28664 Non-provisional of 60/172,411 Dec. 17, 1999 10/100,683 Continuation-in-part of PCT/US00/29363 Oct. 25, 2000 PCT/US00/29363 Non-provisional of 60/215,139 Jun. 30, 2000 PCT/US00/29363 Non-provisional of 60/162,239 Oct. 29, 1999 10/100,683 Continuation-in-part of PCT/US00/29360 Oct. 25, 2000 PCT/US00/29360 Non-provisional of 60/215,138 Jun. 30, 2000 PCT/US00/29360 Non-provisional of 60/162,211 Oct. 29, 1999 10/100,683 Continuation-in-part of PCT/US00/29362 Oct. 25, 2000 PCT/US00/29362 Non-provisional of 60/215,131 Jun. 30, 2000 PCT/US00/29362 Non-provisional of 60/162,240 Oct. 29, 1999 10/100,683 Continuation-in-part of PCT/US00/29365 Oct. 25, 2000 PCT/US00/29365 Non-provisional of 60/219,666 Jul. 21, 2000 PCT/US00/29365 Non-provisional of 60/162,237 Oct. 29, 1999 10/100,683 Continuation-in-part of PCT/US00/29364 Oct. 25, 2000 PCT/US00/29364 Non-provisional of 60/215,134 Jun. 30, 2000 PCT/US00/29364 Non-provisional of 60/162,238 Oct. 29, 1999 10/100,683 Continuation-in-part of PCT/US00/30040 Nov. 01, 2000 PCT/US00/30040 Non-provisional of 60/215,130 Jun. 30, 2000 PCT/US00/30040 Non-provisional of 60/163,580 Nov. 05, 1999 10/100,683 Continuation-in-part of PCT/US00/30037 Nov. 01, 2000 PCT/US00/30037 Non-provisional of 60/215,137 Jun. 30, 2000 PCT/US00/30037 Non-provisional of 60/163,577 Nov. 05, 1999 10/100,683 Continuation-in-part of PCT/US00/30045 Nov. 01, 2000 PCT/US00/30045 Non-provisional of 60/215,133 Jun. 30, 2000 PCT/US00/30045 Non-provisional of 60/163,581 Nov. 05, 1999 10/100,683 Continuation-in-part of PCT/US00/30036 Nov. 01, 2000 PCT/US00/30036 Non-provisional of 60/221,366 Jul. 27, 2000 PCT/US00/30036 Non-provisional of 60/163,576 Nov. 05, 1999 10/100,683 Continuation-in-part of PCT/US00/30039 Nov. 01, 2000 PCT/US00/30039 Non-provisional of 60/221,367 Jul. 27, 2000 PCT/US00/30039 Non-provisional of 60/195,296 Apr. 07, 2000 PCT/US00/30039 Non-provisional of 60/164,344 Nov. 09, 1999 10/100,683 Continuation-in-part of PCT/US00/30654 Nov. 08, 2000 PCT/US00/30654 Non-provisional of 60/221,142 Jul. 27, 2000 PCT/US00/30654 Non-provisional of 60/164,835 Nov. 12, 1999 10/100,683 Continuation-in-part of PCT/US00/30628 Nov. 08, 2000 PCT/US00/30628 Non-provisional of 60/215,140 Jun. 30, 2000 PCT/US00/30628 Non-provisional of 60/164,744 Nov. 12, 1999 10/100,683 Continuation-in-part of PCT/US00/30653 Nov. 08, 2000 PCT/US00/30653 Non-provisional of 60/221,193 Jul. 27, 2000 PCT/US00/30653 Non-provisional of 60/164,735 Nov. 12, 1999 10/100,683 Continuation-in-part of PCT/US00/30629 Nov. 08, 2000 PCT/US00/30629 Non-provisional of 60/222,904 Aug. 03, 2000 PCT/US00/30629 Non-provisional of 60/164,825 Nov. 12, 1999 10/100,683 Continuation-in-part of PCT/US00/30679 Nov. 08, 2000 PCT/US00/30679 Non-provisional of 60/224,007 Aug. 04, 2000 PCT/US00/30679 Non-provisional of 60/164,834 Nov. 12, 1999 10/100,683 Continuation-in-part of PCT/US00/30674 Nov. 08, 2000 PCT/US00/30674 Non-provisional of 60/215,128 Jun. 30, 2000 PCT/US00/30674 Non-provisional of 60/164,750 Nov. 12, 1999 10/100,683 Continuation-in-part of PCT/US00/31162 Nov. 15, 2000 60/215,136 Non-provisional of 60/215,136 Jun. 30, 2000 60/215,136 Non-provisional of 60/166,415 Nov. 19, 1999 10/100,683 Continuation-in-part of PCT/US00/31282 Nov. 15, 2000 PCT/US00/31282 Non-provisional of 60/219,665 Jul. 21, 2000 PCT/US00/31282 Non-provisional of 60/166,414 Nov. 19, 1999 10/100,683 Continuation-in-part of PCT/US00/30657 Nov. 08, 2000 PCT/US00/30657 Non-provisional of 60/215,132 Jun. 30, 2000 PCT/US00/30657 Non-provisional of 60/164,731 Nov. 12, 1999 10/100,683 Continuation-in-part of PCT/US01/01396 Jan. 17, 2001 60/256,968 Non-provisional of 60/256,968 Dec. 21, 2000 60/256,968 Non-provisional of 60/226,280 Aug. 18, 2000 10/100,683 Continuation-in-part of PCT/US01/01387 Jan. 17, 2001 60/259,803 Non-provisional of 60/259,803 Jan. 05, 2001 60/259,803 Non-provisional of 60/226,380 Aug. 18, 2000 10/100,683 Continuation-in-part of PCT/US01/01567 Jan. 17, 2001 PCT/US01/01567 Non-provisional of 60/228,084 Aug. 28, 2000 10/100,683 Continuation-in-part of PCT/US01/01431 Jan. 17, 2001 PCT/US01/01431 Non-provisional of 60/231,968 Sep. 12, 2000 PCT/US01/01431 Continuation-in-part of 09/915,582 Jul. 27, 2001 10/100,683 Continuation-in-part of PCT/US01/01432 Jan. 17, 2001 PCT/US01/01432 Non-provisional of 60/236,326 Sep. 29, 2000 10/100,683 Continuation-in-part of PCT/US01/00544 Jan. 09, 2001 PCT/US01/00544 Non-provisional of 60/234,211 Sep. 20, 2000 10/100,683 Continuation-in-part of PCT/US01/01435 Jan. 17, 2001 PCT/US01/01435 Non-provisional of 60/226,282 Aug. 18, 2000 10/100,683 Continuation-in-part of PCT/US01/01386 Jan. 17, 2001 PCT/US01/01386 Non-provisional of 60/232,104 Sep. 12, 2000 10/100,683 Continuation-in-part of PCT/US01/01565 Jan. 17, 2001 PCT/US01/01565 Non-provisional of 60/234,210 Sep. 20, 2000 10/100,683 Continuation-in-part of PCT/US01/01394 Jan. 17, 2001 PCT/US01/01394 Non-provisional of 60/259,805 Jan. 05, 2001 PCT/US01/01394 Non-provisional of 60/226,278 Aug. 18, 2000 10/100,683 Continuation-in-part of PCT/US01/01434 Jan. 17, 2001 PCT/US01/01434 Non-provisional of 60/259,678 Jan. 05, 2001 PCT/US01/01434 Non-provisional of 60/226,279 Aug. 18, 2000 10/100,683 Continuation-in-part of PCT/US01/01397 Jan. 17, 2001 PCT/US01/01397 Non-provisional of 60/226,281 Aug. 18, 2000 10/100,683 Continuation-in-part of PCT/US01/01385 Jan. 17, 2001 PCT/US01/01385 Non-provisional of 60/231,969 Sep. 12, 2000 10/100,683 Continuation-in-part of PCT/US01/01384 Jan. 17, 2001 PCT/US01/01384 Non-provisional of 60/259,516 Jan. 04, 2001 PCT/US01/01384 Non-provisional of 60/228,086 Aug. 28, 2000 10/100,683 Continuation-in-part of PCT/US01/01383 Jan. 17, 2001 PCT/US01/01383 Non-provisional of 60/259,804 Jan. 05, 2001 PCT/US01/01383 Non-provisional of 60/228,083 Aug. 28, 2000 10/100,683 Continuation-in-part of PCT/US02/05064 Feb. 21, 2002 PCT/US02/05064 Non-provisional of 60/304,444 Jul. 12, 2001 PCT/US02/05064 Non-provisional of 60/270,658 Feb. 23, 2001 10/100,683 Continuation-in-part of PCT/US02/05301 Feb. 21, 2002 PCT/US02/05301 Non-provisional of 60/304,417 Jul. 12, 2001 PCT/US02/05301 Non-provisional of 60/270,625 Feb. 23, 2001 10/100,683 Non-provisional of 60/304,121 Jul. 11, 2001 10/100,683 Non-provisional of 60/295,869 Jun. 06, 2001 10/100,683 Non-provisional of 60/325,209 Sep. 28, 2001 10/100,683 Non-provisional of 60/311,085 Aug. 10, 2001 10/100,683 Non-provisional of 60/330,629 Oct. 26, 2001 10/100,683 Non-provisional of 60/331,046 Nov. 07, 2001 10/100,683 Non-provisional of 60/358,554 Feb. 22, 2002 10/100,683 Non-provisional of 60/358,714 Feb. 25, 2002 ; wherein each of the above applications are all herein incorporated by reference in their entirety.

REFERENCE TO SEQUENCE LISTING AS TEXT FILE

This application refers to a “Sequence Listing” listed below, which is provided as a text file. The text file contains a document entitled “PS902P1D1-SeqList.txt” (3,646,557 bytes, created Jun. 27, 2008), which is hereby incorporated by reference in its entirety.

FIELD OF THE INVENTION

The present invention relates to human secreted proteins/polypeptides, and isolated nucleic acid molecules encoding said proteins/polypeptides, useful for detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune disorders and diseases. Antibodies that bind these polypeptides are also encompassed by the present invention. Also encompassed by the invention are vectors, host cells, and recombinant and synthetic methods for producing said polynucleotides, polypeptides, and/or antibodies. The invention further encompasses screening methods for identifying agonists and antagonists of polynucleotides and polypeptides of the invention. The present invention further encompasses methods and compositions for inhibiting or enhancing the production and function of the polypeptides of the present invention.

BACKGROUND OF THE INVENTION

The immune system is an intricate network of cells, tissues and soluble molecules that function to protect the body from invasion by foreign substances and pathogens. The major cells of the immune system are lymphocytes, including B cells and T cells, and myeloid cells, including basophils, eosinophils, neutrophils, mast cells, monocytes, macrophages and dendritic cells. In addition to these cellular components of the immune system, soluble molecules—such as antibodies, complement proteins, and cytokines—circulate in lymph and blood plasma, and play important roles in immunity.

The immune system can be subdivided into the acquired and innate immune systems. The cells of the innate immune system (e.g., neutrophils, eosinophils, basophils, mast cells) are not antigen specific and their action is not enhanced by repeated exposure to the same antigen. The cells of the acquired immune system (B and T cells) are antigen specific. Repeated exposure of B and T cells to an antigen results in improved immune responses (memory responses) produced by these cell types. The cells and products of the acquired immune system can recruit components of the innate system to mount a focused immune response. For a more extensive review of the immune system, see Fundamental Immunology, 4th edition, Ed. William Paul, Lippincott-Raven Pub. (1998).

An immune response is seldom carried out by a single cell type, but rather requires the coordinated efforts of several cell types. In order to coordinate an immune response, it is necessary that cells of the immune system communicate with each other and with other cells of the body. Communication between cells may be made by cell-cell contact, between membrane bound molecules on each cell, or by the interaction of soluble components of the immune system with cellular receptors. Signaling between cell types may have one or more of a variety of consequences, including activation, proliferation, differentiation, and apoptosis. Activation and differentiation of immune cells may result in the expression or secretion of polypeptides, or other molecules, which in turn affect the function of other cells and/or molecules of the immune system.

Molecules which stimulate or suppress immune system function are known as immunomodulators. These molecules, which include endogenous proteins (e.g., cytokines, cytokine receptors, and intracellular signal transduction molecules), molecules derived from microorganisms, and synthetic agents, may exert their modulatory effects at one or more stages of the immune response, such as antigen recognition, stimulation of cytokine production and release, and/or activation/differentiation of lymphocytes and myeloid cells Immunomodulators may enhance (immunoprophylaxis, immunostimulation), restore (immunosubstitution, immunorestoration) or suppress (immunosuppression, immunodeviation) immunological functions or activities.

Immunomodulatory compounds have many important applications in clinical practice. For example, immunosuppressing agents (which attenuate or prevent unwanted immune responses) can be used to prevent tissue rejection during organ transplantation, to prevent Rh hemolytic disease of the newborn, or to treat autoimmune disorders. A mechanism of action common to many immunosuppressants is the inhibition of T cell activation and/or differentiation. Antilymphocyte antibodies have also been used to attenuate immune system functions. Currently-used immunosuppressive agents can produce a number of side effects which limit their use. Among the most serious secondary effects include kidney and liver toxicity, increased risk of infection, hyperglycemia, neoplasia, and osteoporosis (see, e.g., Freeman, Clin. Biochem. 24(1):9-14 (1991); Mitchison, Dig. Dis. 11(2):78-101 (1993)).

Immunostimulants, which enhance the activity of immune cells and molecules, comprise another class of immunomodulatory agents with important clinical applications. Such applications include, for example, the treatment of immunodeficiency disorders (e.g. AIDS and severe combined immunodeficiency), chronic infectious diseases (e.g. viral hepatitis, papillomavirus, and herpesvirus), and cancer. An important class of endogenous immunostimulants is the cytokines. These soluble signaling molecules are produced by a number of cell types, and are critical to the regulation of the immune response Immunostimulatory mechanisms can include proliferation, differentiation and/or activation of immune cells or progenitors of immune cells. For example, interleukin-2 (IL-2) binds to IL-2 receptors on T lymphocytes and induces proliferation and differentiation. Another cytokine, interferon alpha, stimulates the immune system through a variety of mechanisms, including activation of macrophages, T lymphocytes, and natural killer cells. Interferon alpha also induces the expression of antiviral proteins (see Chapter 50, The Pharmacological Basis of Therapeutics, 9^(th) Edition, Eds. Hardman, Limbird, Molinoff, Ruddon, and Gilman, McGraw Hill (1996)). Limitations of current immunostimulant therapies include anaphylaxis, pulmonary edema, and renal toxicity, to name a few.

The discovery of new human immune related polynucleotides, the polypeptides encoded by them, and antibodies that immunospecifically bind these polypeptides, satisfies a need in the art by providing new compositions which are useful in the diagnosis, treatment, prevention and/or prognosis of disorders of the immune system, including, but not limited to, autoimmune disorders (e.g., systemic lupus erythematosus, rheumatoid arthritis, idiopathic thrombocytopenic purpura and multiple sclerosis), immunodeficiencies (e.g., X-linked agammaglobulinemia, severe combined immunodeficiency, Wiskott-Aldrich syndrome, and ataxia telangiectasia), chronic infections (e.g., HIV, viral hepatitis, and herpesvirus), and neoplastic disorders. See, e.g. “Immune Activity” section infra. Additionally, immune related molecules would be useful as agents to boost immune responsiveness to pathogens or to suppress immune reactions, for example as is necessary in conjunction with organ transplantation.

SUMMARY OF THE INVENTION

The present invention relates to novel secreted proteins. More specifically, isolated nucleic acid molecules are provided encoding novel secreted polypeptides. Novel polypeptides and antibodies that bind to these polypeptides are provided. Also provided are vectors, host cells, and recombinant and synthetic methods for producing human polynucleotides, polypeptides, and/or antibodies. The invention further relates to diagnostic and therapeutic methods useful for detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune diseases and disorders. The invention further relates to screening methods for identifying agonists and antagonists of polynucleotides and polypeptides of the invention. The present invention further relates to methods and/or compositions for inhibiting or enhancing the production and function of the polypeptides of the present invention.

DETAILED DESCRIPTION Polynucleotides and Polypeptides of the Invention Description of Table 1A

Table 1A summarizes information concerning certain polynucleotides and polypeptides of the invention. The first column provides the gene number in the application for each clone identifier. The second column provides a unique clone identifier, “Clone ID:”, for a cDNA clone related to each contig sequence disclosed in Table 1A. Third column, the cDNA Clones identified in the second column were deposited as indicated in the third column (i.e. by ATCC™ Deposit No:Z and deposit date). Some of the deposits contain multiple different clones corresponding to the same gene. In the fourth column, “Vector” refers to the type of vector contained in the corresponding cDNA Clone identified in the second column. In the fifth column, the nucleotide sequence identified as “NT SEQ ID NO:X” was assembled from partially homologous (“overlapping”) sequences obtained from the corresponding cDNA clone identified in the second column and, in some cases, from additional related cDNA clones. The overlapping sequences were assembled into a single contiguous sequence of high redundancy (usually three to five overlapping sequences at each nucleotide position), resulting in a final sequence identified as SEQ ID NO:X. In the sixth column, “Total NT Seq.” refers to the total number of nucleotides in the contig sequence identified as SEQ ID NO:X.” The deposited clone may contain all or most of these sequences, reflected by the nucleotide position indicated as “5′ NT of Clone Seq.” (seventh column) and the “3′ NT of Clone Seq.” (eighth column) of SEQ ID NO:X. In the ninth column, the nucleotide position of SEQ ID NO:X of the putative start codon (methionine) is identified as “5′ NT of Start Codon.” Similarly, in column ten, the nucleotide position of SEQ ID NO:X of the predicted signal sequence is identified as “5′ NT of First AA of Signal Pep.” In the eleventh column, the translated amino acid sequence, beginning with the methionine, is identified as “AA SEQ ID NO:Y,” although other reading frames can also be routinely translated using known molecular biology techniques. The polypeptides produced by these alternative open reading frames are specifically contemplated by the present invention.

In the twelfth and thirteenth columns of Table 1A, the first and last amino acid position of SEQ ID NO:Y of the predicted signal peptide is identified as “First AA of Sig Pep” and “Last AA of Sig Pep.” In the fourteenth column, the predicted first amino acid position of SEQ ID NO:Y of the secreted portion is identified as “Predicted First AA of Secreted Portion”. The amino acid position of SEQ ID NO:Y of the last amino acid encoded by the open reading frame is identified in the fifteenth column as “Last AA of ORF”.

SEQ ID NO:X (where X may be any of the polynucleotide sequences disclosed in the sequence listing) and the translated SEQ ID NO:Y (where Y may be any of the polypeptide sequences disclosed in the sequence listing) are sufficiently accurate and otherwise suitable for a variety of uses well known in the art and described further below. For instance, SEQ ID NO:X is useful for designing nucleic acid hybridization probes that will detect nucleic acid sequences contained in SEQ ID NO:X or the cDNA contained in the deposited clone. These probes will also hybridize to nucleic acid molecules in biological samples, thereby enabling a variety of forensic and diagnostic methods of the invention. Similarly, polypeptides identified from SEQ ID NO:Y may be used, for example, to generate antibodies which bind specifically to proteins containing the polypeptides and the secreted proteins encoded by the cDNA clones identified in Table 1A and/or elsewhere herein

Nevertheless, DNA sequences generated by sequencing reactions can contain sequencing errors. The errors exist as misidentified nucleotides, or as insertions or deletions of nucleotides in the generated DNA sequence. The erroneously inserted or deleted nucleotides cause frame shifts in the reading frames of the predicted amino acid sequence. In these cases, the predicted amino acid sequence diverges from the actual amino acid sequence, even though the generated DNA sequence may be greater than 99.9% identical to the actual DNA sequence (for example, one base insertion or deletion in an open reading frame of over 1000 bases).

Accordingly, for those applications requiring precision in the nucleotide sequence or the amino acid sequence, the present invention provides not only the generated nucleotide sequence identified as SEQ ID NO:X, and the predicted translated amino acid sequence identified as SEQ ID NO:Y, but also a sample of plasmid DNA containing a human cDNA of the invention deposited with the ATCC™, as set forth in Table 1A. The nucleotide sequence of each deposited plasmid can readily be determined by sequencing the deposited plasmid in accordance with known methods

The predicted amino acid sequence can then be verified from such deposits. Moreover, the amino acid sequence of the protein encoded by a particular plasmid can also be directly determined by peptide sequencing or by expressing the protein in a suitable host cell containing the deposited human cDNA, collecting the protein, and determining its sequence.

Also provided in Table 1A is the name of the vector which contains the cDNA plasmid. Each vector is routinely used in the art. The following additional information is provided for convenience.

Vectors Lambda Zap (U.S. Pat. Nos. 5,128,256 and 5,286,636), Uni-Zap XR (U.S. Pat. Nos. 5,128,256 and 5,286,636), Zap Express (U.S. Pat. Nos. 5,128,256 and 5,286,636), pBLUESCRIPT™ (pBS) (Short, J. M. et al., Nucleic Acids Res. 16:7583-7600 (1988); Alting-Mees, M. A. and Short, J. M., Nucleic Acids Res. 17:9494 (1989)) and pBK (Alting-Mees, M. A. et al., Strategies 5:58-61 (1992)) are commercially available from STRATAGENE™ Cloning Systems, Inc., 11011 N. Torrey Pines Road, La Jolla, Calif., 92037. pBS contains an ampicillin resistance gene and pBK contains a neomycin resistance gene. Phagemid pBS may be excised from the Lambda Zap and Uni-Zap XR vectors, and phagemid pBK may be excised from the Zap Express vector. Both phagemids may be transformed into E. coli strain XL-1 Blue, also available from STRATAGENET™.

Vectors pSport1, pCMVSport 1.0, pCMVSport 2.0 and pCMVSport 3.0, were obtained from Life Technologies, Inc., P.O. Box 6009, Gaithersburg, Md. 20897. All Sport vectors contain an ampicillin resistance gene and may be transformed into E. coli strain DH10B, also available from Life Technologies. See, for instance, Gruber, C. E., et al., Focus 15:59 (1993). Vector lafmid BA (Bento Soares, Columbia University, New York, N.Y.) contains an ampicillin resistance gene and can be transformed into E. coli strain XL-1 Blue. Vector pCR®2.1, which is available from Invitrogen, 1600 Faraday Avenue, Carlsbad, Calif. 92008, contains an ampicillin resistance gene and may be transformed into E. coli strain DH10B, available from Life Technologies. See, for instance, Clark, J. M., Nuc. Acids Res. 16:9677-9686 (1988) and Mead, D. et al., Bio/Technology 9: (1991).

The present invention also relates to the genes corresponding to SEQ ID NO:X, SEQ ID NO:Y, and/or a deposited cDNA (cDNA Clone ID). The corresponding gene can be isolated in accordance with known methods using the sequence information disclosed herein. Such methods include, but are not limited to, preparing probes or primers from the disclosed sequence and identifying or amplifying the corresponding gene from appropriate sources of genomic material.

Also provided in the present invention are allelic variants, orthologs, and/or species homologs. Procedures known in the art can be used to obtain full-length genes, allelic variants, splice variants, full-length coding portions, orthologs, and/or species homologs of genes corresponding to SEQ ID NO:X and SEQ ID NO:Y using information from the sequences disclosed herein or the clones deposited with the ATCC™. For example, allelic variants and/or species homologs may be isolated and identified by making suitable probes or primers from the sequences provided herein and screening a suitable nucleic acid source for allelic variants and/or the desired homologue.

The present invention provides a polynucleotide comprising, or alternatively consisting of, the nucleic acid sequence of SEQ ID NO:X and/or a cDNA contained in ATCC™ Deposit No.Z. The present invention also provides a polypeptide comprising, or alternatively, consisting of, the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X, and/or a polypeptide encoded by a cDNA contained in ATCC™ deposit No.Z. Polynucleotides encoding a polypeptide comprising, or alternatively consisting of the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X and/or a polypeptide encoded by the cDNA contained in ATCC™ Deposit No.Z, are also encompassed by the invention. The present invention further encompasses a polynucleotide comprising, or alternatively consisting of the complement of the nucleic acid sequence of SEQ ID NO:X, and/or the complement of the coding strand of the cDNA contained in ATCC™ Deposit No.Z.

Description of Table 1B (Comprised of Tables 1B.1 and 1B.2)

Table 1B.1 and Table 1B.2 summarize some of the polynucleotides encompassed by the invention (including cDNA clones related to the sequences (Clone ID:), contig sequences (contig identifier (Contig ID:) and contig nucleotide sequence identifiers (SEQ ID NO:X)) and further summarizes certain characteristics of these polynucleotides and the polypeptides encoded thereby. The first column of Tables 1B.1 and 1B.2 provide the gene numbers in the application for each clone identifier. The second column of Tables 1B.1 and 1B.2 provide unique clone identifiers, “Clone ID:”, for cDNA clones related to each contig sequence disclosed in Table 1A and/or Table 1B. The third column of Tables 1B.1 and 1B.2 provide unique contig identifiers, “Contig ID:” for each of the contig sequences disclosed in these tables. The fourth column of Tables 1B.1 and 1B.2 provide the sequence identifiers, “SEQ ID NO:X”, for each of the contig sequences disclosed in Table 1A and/or 1B.

Table 1B.1

The fifth column of Table 1B.1, “ORF (From-To)”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence of SEQ ID NO:X that delineates the preferred open reading frame (ORF) that encodes the amino acid sequence shown in the sequence listing and referenced in Table 1B.1 as SEQ ID NO:Y (column 6). Column 7 of Table 1B.1 lists residues comprising predicted epitopes contained in the polypeptides encoded by each of the preferred ORFs (SEQ ID NO:Y). Identification of potential immunogenic regions was performed according to the method of Jameson and Wolf (CABIOS, 4; 181-186 (1988)); specifically, the Genetics Computer Group (GCG) implementation of this algorithm, embodied in the program PEPTIDESTRUCTURE (Wisconsin Package v10.0, Genetics Computer Group (GCG), Madison, Wisc.). This method returns a measure of the probability that a given residue is found on the surface of the protein. Regions where the antigenic index score is greater than 0.9 over at least 6 amino acids are indicated in Table 1B.1 as “Predicted Epitopes”. In particular embodiments, polypeptides of the invention comprise, or alternatively consist of, one, two, three, four, five or more of the predicted epitopes described in Table 1B.1. It will be appreciated that depending on the analytical criteria used to predict antigenic determinants, the exact address of the determinant may vary slightly. Column 8 of Table 1B.1 (“Cytologic Band”) provides the chromosomal location of polynucleotides corresponding to SEQ ID NO:X. Chromosomal location was determined by finding exact matches to EST and cDNA sequences contained in the NCBI (National Center for Biotechnology Information) UniGene database. Given a presumptive chromosomal location, disease locus association was determined by comparison with the Morbid Map, derived from Online Mendelian Inheritance in Man (Online Mendelian Inheritance in Man, OMIM™. McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University (Baltimore, Md.) and National Center for Biotechnology Information, National Library of Medicine (Bethesda, Md.) 2000. If the putative chromosomal location of the Query overlaps with the chromosomal location of a Morbid Map entry, an OMIM identification number is disclosed in Table 1B.1, column 9 labeled “OMIM Disease Reference(s)”. A key to the OMIM reference identification numbers is provided in Table 5.

Table 1B.2

Column 5 of Table 1B.2, “Tissue Distribution” shows the expression profile of tissue, cells, and/or cell line libraries which express the polynucleotides of the invention. The first code number shown in Table 1B.2 column 5 (preceding the colon), represents the tissue/cell source identifier code corresponding to the key provided in Table 4. Expression of these polynucleotides was not observed in the other tissues and/or cell libraries tested. The second number in column 5 (following the colon), represents the number of times a sequence corresponding to the reference polynucleotide sequence (e.g., SEQ ID NO:X) was identified in the corresponding tissue/cell source. Those tissue/cell source identifier codes in which the first two letters are “AR” designate information generated using DNA array technology. Utilizing this technology, cDNAs were amplified by PCR and then transferred, in duplicate, onto the array. Gene expression was assayed through hybridization of first strand cDNA probes to the DNA array. cDNA probes were generated from total RNA extracted from a variety of different tissues and cell lines. Probe synthesis was performed in the presence of ³³P dCTP, using oligo(dT) to prime reverse transcription. After hybridization, high stringency washing conditions were employed to remove non-specific hybrids from the array. The remaining signal, emanating from each gene target, was measured using a Phosphorimager. Gene expression was reported as Phosphor Stimulating Luminescence (PSL) which reflects the level of phosphor signal generated from the probe hybridized to each of the gene targets represented on the array. A local background signal subtraction was performed before the total signal generated from each array was used to normalize gene expression between the different hybridizations. The value presented after “[array code]:” represents the mean of the duplicate values, following background subtraction and probe normalization. One of skill in the art could routinely use this information to identify normal and/or diseased tissue(s) which show a predominant expression pattern of the corresponding polynucleotide of the invention or to identify polynucleotides which show predominant and/or specific tissue and/or cell expression.

Description of Table 1C

Table 1C summarizes additional polynucleotides encompassed by the invention (including cDNA clones related to the sequences (Clone ID:), contig sequences (contig identifier (Contig ID:) contig nucleotide sequence identifiers (SEQ ID NO:X)), and genomic sequences (SEQ ID NO:B). The first column provides a unique clone identifier, “Clone ID:”, for a cDNA clone related to each contig sequence. The second column provides the sequence identifier, “SEQ ID NO:X”, for each contig sequence. The third column provides a unique contig identifier, “Contig ID:” for each contig sequence. The fourth column, provides a BAC identifier “BAC ID NO:A” for the BAC clone referenced in the corresponding row of the table. The fifth column provides the nucleotide sequence identifier, “SEQ ID NO:B” for a fragment of the BAC clone identified in column four of the corresponding row of the table. The sixth column, “Exon From-To”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence of SEQ ID NO:B which delineate certain polynucleotides of the invention that are also exemplary members of polynucleotide sequences that encode polypeptides of the invention (e.g., polypeptides containing amino acid sequences encoded by the polynucleotide sequences delineated in column six, and fragments and variants thereof).

Description of Tables 1D.1 and 1D.2

Tables 1D.1 and 1D.2: In preferred embodiments, the present invention encompasses a method of detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune diseases or disorders; comprising administering to a patient in which such treatment, prevention, or amelioration is desired a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof) represented by Table 1A, Table 1B, and Table 1C, in an amount effective to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate the disease or disorder.

As indicated in Tables 1D.1 and 1D.2, the polynucleotides, polypeptides, agonists, or antagonists of the present invention (including antibodies) can be used in assays to test for one or more biological activities. If these polynucleotides and polypeptides do exhibit activity in a particular assay, it is likely that these molecules may be involved in the diseases associated with the biological activity. Thus, the polynucleotides or polypeptides, or agonists or antagonists thereof (including antibodies) could be used to treat the associated disease.

Tables 1D.1 and 1D.2 provide information related to biological activities for polynucleotides and polypeptides of the invention (including antibodies, agonists, and/or antagonists thereof). Tables 1D.1 and 1D.2 also provide information related to assays which may be used to test polynucleotides and polypeptides of the invention (including antibodies, agonists, and/or antagonists thereof) for the corresponding biological activities. The first column of Table 1D.1 (“Gene No.”) provides the gene number in the application for each clone identifier. The second column of Table 1D.1 (“cDNA Clone ID:”) provides the unique clone identifier for each clone as previously described and indicated in Tables 1A, 1B, and 1C. The third column of Table 1D.1 (“AA SEQ ID NO:Y”) indicates the Sequence Listing SEQ ID Number for polypeptide sequences encoded by the corresponding cDNA clones (also as indicated in Tables 1A, 1B, and 2). The fourth column of Table 1D.1 (“Biological Activity”) indicates a biological activity corresponding to the indicated polypeptides (or polynucleotides encoding said polypeptides). The first column of Table 1D.2 (“Biological Activity No.”) provides the biological activity number corresponding to each biological activity recited in the fourth column of Table 1D.1. The second column of Table 1D.2 (“Biological Activity”) indicates the biological activity corresponding to each biological activity number (also as indicated in Table 1D.1). The third column of Table 1D.2 (“Exemplary Activity Assay”) further describes the corresponding biological activity and provides information pertaining to the various types of assays which may be performed to test, demonstrate, or quantify the corresponding biological activity. Tables 1D.1 and 1D.2 describes the use of FMAT technology, inter alia, for testing or demonstrating various biological activities. Fluorometric microvolume assay technology (FMAT) is a fluorescence-based system that provides a means to perform nonradioactive cell- and bead-based assays to detect activation of cell signal transduction pathways. This technology was designed specifically for ligand binding and immunological assays. Using this technology, fluorescent cells or beads at the bottom of the well are detected as localized areas of concentrated fluorescence using a data processing system. Unbound fluorophore comprising the background signal is ignored, allowing for a wide variety of homogeneous assays. FMAT technology may be used for peptide ligand binding assays, immunofluorescence, apoptosis, cytotoxicity, and bead-based immunocapture assays. See, Miraglia S et. al., “Homogeneous cell and bead based assays for high throughput screening using fluorometric microvolume assay technology,” Journal of Biomolecular Screening; 4:193-204 (1999). In particular, FMAT technology may be used to test, confirm, and/or identify the ability of polypeptides (including polypeptide fragments and variants) to activate signal transduction pathways. For example, FMAT technology may be used to test, confirm, and/or identify the ability of polypeptides to upregulate production of immunomodulatory proteins (such as, for example, interleukins, GM-CSF, Rantes, and Tumor Necrosis factors, as well as other cellular regulators (e.g. insulin)).

Tables 1D.1 and 1D.2 also describe the use of kinase assays for testing, demonstrating, or quantifying biological activity. In this regard, the phosphorylation and de-phosphorylation of specific amino acid residues (e.g. Tyrosine, Serine, Threonine) on cell-signal transduction proteins provides a fast, reversible means for activation and de-activation of cellular signal transduction pathways. Moreover, cell signal transduction via phosphorylation/de-phosphorylation is crucial to the regulation of a wide variety of cellular processes (e.g. proliferation, differentiation, migration, apoptosis, etc.). Accordingly, kinase assays provide a powerful tool useful for testing, confirming, and/or identifying polypeptides (including polypeptide fragments and variants) that mediate cell signal transduction events via protein phosphorylation. See e.g., Forrer, P., Tamaskovic R., and Jaussi, R. “Enzyme-Linked Immunosorbent Assay for Measurement of INK, ERK, and p38 Kinase Activities” Biol. Chem. 379(8-9): 1101-1110 (1998).

Description of Table 1E

Polynucleotides encoding polypeptides of the present invention can be used in assays to test for one or more biological activities. One such biological activity which may be tested includes the ability of polynucleotides and polypeptides of the invention to stimulate up-regulation or down-regulation of expression of particular genes and proteins. Hence, if polynucleotides and polypeptides of the present invention exhibit activity in altering particular gene and protein expression patterns, it is likely that these polynucleotides and polypeptides of the present invention may be involved in, or capable of effecting changes in, diseases associated with the altered gene and protein expression profiles. Hence, polynucleotides, polypeptides, or antibodies of the present invention could be used to treat said associated diseases.

TaqMan® assays may be performed to assess the ability of polynucleotides (and polypeptides they encode) to alter the expression pattern of particular “target” genes. TaqMan® reactions are performed to evaluate the ability of a test agent to induce or repress expression of specific genes in different cell types. TaqMan® gene expression quantification assays (“TaqMan® assays”) are well known to, and routinely performed by, those of ordinary skill in the art. TaqMan® assays are performed in a two step reverse transcription/polymerase chain reaction (RT-PCR). In the first (RT) step, cDNA is reverse transcribed from total RNA samples using random hexamer primers. In the second (PCR) step, PCR products are synthesized from the cDNA using gene specific primers.

To quantify gene expression the Taqman® PCR reaction exploits the 5′ nuclease activity of AmpliTaq Gold® DNA Polymerase to cleave a Taqman® probe (distinct from the primers) during PCR. The Taqman® probe contains a reporter dye at the 5′-end of the probe and a quencher dye at the 3′ end of the probe. When the probe is intact, the proximity of the reporter dye to the quencher dye results in suppression of the reporter fluorescence. During PCR, if the target of interest is present, the probe specifically anneals between the forward and reverse primer sites. AmpliTaq Fold DNA Polymerase then cleaves the probe between the reporter and quencher when the probe hybridizes to the target, resulting in increased fluorescence of the reporter (see FIG. 2). Accumulation of PCR products is detected directly by monitoring the increase in fluorescence of the reporter dye.

After the probe fragments are displaced from the target, polymerization of the strand continues. The 3′-end of the probe is blocked to prevent extension of the probe during PCR. This process occurs in every cycle and does not interfere with the exponential accumulation of product. The increase in fluorescence signal is detected only if the target sequence is complementary to the probe and is amplified during PCR. Because of these requirements, any nonspecific amplification is not detected.

For test sample preparation, vector controls or constructs containing the coding sequence for the gene of interest are transfected into cells, such as for example 293T cells, and supernatants collected after 48 hours. For cell treatment and RNA isolation, multiple primary human cells or human cell lines are used; such cells may include but are not limited to, Normal Human Dermal Fibroblasts, Aortic Smooth Muscle, Human Umbilical Vein Endothelial Cells, HepG2, Daudi, Jurkat, U937, Caco, and THP-1 cell lines. Cells are plated in growth media and growth is arrested by culturing without media change for 3 days, or by switching cells to low serum media and incubating overnight. Cells are treated for 1, 6, or 24 hours with either vector control supernatant or sample supernatant (or purified/partially purified protein preparations in buffer). Total RNA is isolated; for example, by using Trizol extraction or by using the Ambion RNAqueous™-4PCR RNA isolation system. Expression levels of multiple genes are analyzed using TAQMAN, and expression in the test sample is compared to control vector samples to identify genes induced or repressed. Each of the above described techniques are well known to, and routinely performed by, those of ordinary skill in the art.

Table 1E indicates particular disease classes and preferred indications for which polynucleotides, polypeptides, or antibodies of the present invention may be used in detecting, diagnosing, preventing, treating and/or ameliorating said diseases and disorders based on “target” gene expression patterns which may be up- or down-regulated by polynucleotides (and the encoded polypeptides) corresponding to each indicated cDNA Clone ID (shown in Table 1E, Column 2).

Thus, in preferred embodiments, the present invention encompasses a method of detecting, diagnosing, preventing, treating, and/or ameliorating a disease or disorder listed in the “Disease Class” and/or “Preferred Indication” columns of Table 1E; comprising administering to a patient in which such detection, diagnosis, prevention, or treatment is desired a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof) in an amount effective to detect, diagnose, prevent, treat, or ameliorate the disease or disorder. The first and second columns of Table 1D.1 show the “Gene No.” and “cDNA Clone ID No.”, respectively, indicating certain nucleic acids and proteins (or antibodies against the same) of the invention (including polynucleotide, polypeptide, and antibody fragments or variants thereof) that may be used in detecting, diagnosing, preventing, treating, or ameliorating the disease(s) or disorder(s) indicated in the corresponding row in the “Disease Class” or “Preferred Indication” Columns of Table 1E.

In another embodiment, the present invention also encompasses methods of detecting, diagnosing, preventing, treating, or ameliorating a disease or disorder listed in the “Disease Class” or “Preferred Indication” Columns of Table 1E; comprising administering to a patient combinations of the proteins, nucleic acids, or antibodies of the invention (or fragments or variants thereof), sharing similar indications as shown in the corresponding rows in the “Disease Class” or “Preferred Indication” Columns of Table 1E.

The “Disease Class” Column of Table 1E provides a categorized descriptive heading for diseases, disorders, and/or conditions (more fully described below) that may be detected, diagnosed, prevented, treated, or ameliorated by a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof).

The “Preferred Indication” Column of Table 1E describes diseases, disorders, and/or conditions that may be detected, diagnosed, prevented, treated, or ameliorated by a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof).

The “Cell Line” and “Exemplary Targets” Columns of Table 1E indicate particular cell lines and target genes, respectively, which may show altered gene expression patterns (i.e., up- or down-regulation of the indicated target gene) in Taqman assays, performed as described above, utilizing polynucleotides of the cDNA Clone ID shown in the corresponding row. Alteration of expression patterns of the indicated “Exemplary Target” genes is correlated with a particular “Disease Class” and/or “Preferred Indication” as shown in the corresponding row under the respective column headings.

The “Exemplary Accessions” Column indicates GenBank Accessions (available online through the National Center for Biotechnology Information (NCBI)) which correspond to the “Exemplary Targets” shown in the adjacent row.

The recitation of “Cancer” in the “Disease Class” Column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof) may be used for example, to detect, diagnose, prevent, treat, and/or ameliorate neoplastic diseases and/or disorders (e.g., leukemias, cancers, etc., as described below under “Hyperproliferative Disorders”).

The recitation of “Immune” in the “Disease Class” column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof), may be used for example, to detect, diagnose, prevent, treat, and/or ameliorate diseases and/or disorders relating to neoplastic diseases (e.g., as described below under “Hyperproliferative Disorders”), blood disorders (e.g., as described below under “Immune Activity” “Cardiovascular Disorders” and/or “Blood-Related Disorders”), and infections (e.g., as described below under “Infectious Disease”).

The recitation of “Angiogenesis” in the “Disease Class” column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof), may be used for example, to detect, diagnose, treat, prevent, and/or ameliorate diseases and/or disorders relating to neoplastic diseases (e.g., as described below under “Hyperproliferative Disorders”), diseases and/or disorders of the cardiovascular system (e.g., as described below under “Cardiovascular Disorders”), diseases and/or disorders involving cellular and genetic abnormalities (e.g., as described below under “Diseases at the Cellular Level”), diseases and/or disorders involving angiogenesis (e.g., as described below under “Anti-Angiogenesis Activity”), to promote or inhibit cell or tissue regeneration (e.g., as described below under “Regeneration”), or to promote wound healing (e.g., as described below under “Wound Healing and Epithelial Cell Proliferation”).

The recitation of “Diabetes” in the “Disease Class” column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof), may be used for example, to detect, diagnose, treat, prevent, and/or ameliorate diabetes (including diabetes mellitus types I and II), as well as diseases and/or disorders associated with, or consequential to, diabetes (e.g. as described below under “Endocrine Disorders,” “Renal Disorders,” and “Gastrointestinal Disorders”).

Description of Table 2

Table 2 summarizes homology and features of some of the polypeptides of the invention. The first column provides a unique clone identifier, “Clone ID:”, corresponding to a cDNA clone disclosed in Table 1A or Table 1B. The second column provides the unique contig identifier, “Contig ID:” corresponding to contigs in Table 1B and allowing for correlation with the information in Table 1B. The third column provides the sequence identifier, “SEQ ID NO:X”, for the contig polynucleotide sequence. The fourth column provides the analysis method by which the homology/identity disclosed in the Table was determined. Comparisons were made between polypeptides encoded by the polynucleotides of the invention and either a non-redundant protein database (herein referred to as “NR”), or a database of protein families (herein referred to as “PFAM”) as further described below. The fifth column provides a description of the PFAM/NR hit having a significant match to a polypeptide of the invention. Column six provides the accession number of the PFAM/NR hit disclosed in the fifth column. Column seven, “Score/Percent Identity”, provides a quality score or the percent identity, of the hit disclosed in columns five and six. Columns 8 and 9, “NT From” and “NT To” respectively, delineate the polynucleotides in “SEQ ID NO:X” that encode a polypeptide having a significant match to the PFAM/NR database as disclosed in the fifth and sixth columns. In specific embodiments polypeptides of the invention comprise, or alternatively consist of, an amino acid sequence encoded by a polynucleotide in SEQ ID NO:X as delineated in columns 8 and 9, or fragments or variants thereof.

Description of Table 3

Table 3 provides polynucleotide sequences that may be disclaimed according to certain embodiments of the invention. The first column provides a unique clone identifier, “Clone ID”, for a cDNA clone related to contig sequences disclosed in Table 1B. The second column provides the sequence identifier, “SEQ ID NO:X”, for contig sequences disclosed in Table 1A and/or Table 1B. The third column provides the unique contig identifier, “Contig ID:”, for contigs disclosed in Table 1B. The fourth column provides a unique integer ‘a’ where ‘a’ is any integer between 1 and the final nucleotide minus 15 of SEQ ID NO:X, and the fifth column provides a unique integer ‘b’ where ‘b’ is any integer between 15 and the final nucleotide of SEQ ID NO:X, where both a and b correspond to the positions of nucleotide residues shown in SEQ ID NO:X, and where b is greater than or equal to a +14. For each of the polynucleotides shown as SEQ ID NO:X, the uniquely defined integers can be substituted into the general formula of a-b, and used to describe polynucleotides which may be preferably excluded from the invention. In certain embodiments, preferably excluded from the invention are at least one, two, three, four, five, ten, or more of the polynucleotide sequence(s) having the accession number(s) disclosed in the sixth column of this Table (including for example, published sequence in connection with a particular BAC clone). In further embodiments, preferably excluded from the invention are the specific polynucleotide sequence(s) contained in the clones corresponding to at least one, two, three, four, five, ten, or more of the available material having the accession numbers identified in the sixth column of this Table (including for example, the actual sequence contained in an identified BAC clone).

Description of Table 4

Table 4 provides a key to the tissue/cell source identifier code disclosed in Table 1B.2, column 5 Column 1 provides the tissue/cell source identifier code disclosed in Table 1B.2, Column 5. Columns 2-5 provide a description of the tissue or cell source. Note that “Description” and “Tissue” sources (i.e. columns 2 and 3) having the prefix “a_” indicates organs, tissues, or cells derived from “adult” sources. Codes corresponding to diseased tissues are indicated in column 6 with the word “disease.” The use of the word “disease” in column 6 is non-limiting. The tissue or cell source may be specific (e.g. a neoplasm), or may be disease-associated (e.g., a tissue sample from a normal portion of a diseased organ). Furthermore, tissues and/or cells lacking the “disease” designation may still be derived from sources directly or indirectly involved in a disease state or disorder, and therefore may have a further utility in that disease state or disorder. In numerous cases where the tissue/cell source is a library, column 7 identifies the vector used to generate the library.

Description of Table 5

Table 5 provides a key to the OMIM reference identification numbers disclosed in Table 1B.1, column 9. OMIM reference identification numbers (Column 1) were derived from Online Mendelian Inheritance in Man (Online Mendelian Inheritance in Man, OMIM. McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University (Baltimore, Md.) and National Center for Biotechnology Information, National Library of Medicine, (Bethesda, Md.) 2000. Column 2 provides diseases associated with the cytologic band disclosed in Table 1B.1, column 8, as determined using the Morbid Map database.

Description of Table 6

Table 6 summarizes some of the ATCC™ Deposits, Deposit dates, and ATCC™ designation numbers of deposits made with the ATCC™ in connection with the present application. These deposits were made in addition to those described in the Table 1A.

Description of Table 7

Table 7 shows the cDNA libraries sequenced, and ATCC™ designation numbers and vector information relating to these cDNA libraries.

The first column shows the first four letters indicating the Library from which each library clone was derived. The second column indicates the catalogued tissue description for the corresponding libraries. The third column indicates the vector containing the corresponding clones. The fourth column shows the ATCC™ deposit designation for each library clone as indicated by the deposit information in Table 6.

Definitions

The following definitions are provided to facilitate understanding of certain terms used throughout this specification.

In the present invention, “isolated” refers to material removed from its original environment (e.g., the natural environment if it is naturally occurring), and thus is altered “by the hand of man” from its natural state. For example, an isolated polynucleotide could be part of a vector or a composition of matter, or could be contained within a cell, and still be “isolated” because that vector, composition of matter, or particular cell is not the original environment of the polynucleotide. The term “isolated” does not refer to genomic or cDNA libraries, whole cell total or mRNA preparations, genomic DNA preparations (including those separated by electrophoresis and transferred onto blots), sheared whole cell genomic DNA preparations or other compositions where the art demonstrates no distinguishing features of the polynucleotide/sequences of the present invention.

In the present invention, a “secreted” protein refers to those proteins capable of being directed to the ER, secretory vesicles, or the extracellular space as a result of a signal sequence, as well as those proteins released into the extracellular space without necessarily containing a signal sequence. If the secreted protein is released into the extracellular space, the secreted protein can undergo extracellular processing to produce a “mature” protein. Release into the extracellular space can occur by many mechanisms, including exocytosis and proteolytic cleavage.

As used herein, a “polynucleotide” refers to a molecule having a nucleic acid sequence encoding SEQ ID NO:Y or a fragment or variant thereof (e.g., the polypeptide delineated in columns fourteen and fifteen of Table 1A); a nucleic acid sequence contained in SEQ ID NO:X (as described in column 5 of Table 1A and/or Table 1B) or the complement thereof; a cDNA sequence contained in Clone ID: (as described in column 2 of Table 1A and/or Table 1B and contained within a library deposited with the ATCC™); a nucleotide sequence encoding the polypeptide encoded by a nucleotide sequence in SEQ ID NO:B as defined in column 6 (EXON From-To) of Table 1C or a fragment or variant thereof; or a nucleotide coding sequence in SEQ ID NO:B as defined in column 6 of Table 1C or the complement thereof. For example, the polynucleotide can contain the nucleotide sequence of the full length cDNA sequence, including the 5′ and 3′ untranslated sequences, the coding region, as well as fragments, epitopes, domains, and variants of the nucleic acid sequence. Moreover, as used herein, a “polypeptide” refers to a molecule having an amino acid sequence encoded by a polynucleotide of the invention as broadly defined (obviously excluding poly-Phenylalanine or poly-Lysine peptide sequences which result from translation of a polyA tail of a sequence corresponding to a cDNA).

In the present invention, “SEQ ID NO:X” was often generated by overlapping sequences contained in multiple clones (contig analysis). A representative clone containing all or most of the sequence for SEQ ID NO:X is deposited at Human Genome Sciences, Inc. (HGS) in a catalogued and archived library. As shown, for example, in column 2 of Table 1B, each clone is identified by a cDNA Clone ID (identifier generally referred to herein as Clone ID:). Each Clone ID is unique to an individual clone and the Clone ID is all the information needed to retrieve a given clone from the HGS library. Table 7 provides a list of the deposited cDNA libraries. One can use the Clone ID: to determine the library source by reference to Tables 6 and 7. Table 7 lists the deposited cDNA libraries by name and links each library to an ATCC™ Deposit. Library names contain four characters, for example, “HTWE.” The name of a cDNA clone (Clone ID) isolated from that library begins with the same four characters, for example “HTWEP07”. As mentioned below, Table 1A and/or Table 1B correlates the Clone ID names with SEQ ID NO:X. Thus, starting with an SEQ ID NO:X, one can use Tables 1A, 1B, 6, 7, and 9 to determine the corresponding Clone ID, which library it came from and which ATCC™ deposit the library is contained in. Furthermore, it is possible to retrieve a given cDNA clone from the source library by techniques known in the art and described elsewhere herein. The ATCC™ is located at 10801 University Boulevard, Manassas, Va. 20110-2209, USA. The ATCC™ deposits were made pursuant to the terms of the Budapest Treaty on the international recognition of the deposit of microorganisms for the purposes of patent procedure.

In specific embodiments, the polynucleotides of the invention are at least 15, at least 30, at least 50, at least 100, at least 125, at least 500, or at least 1000 continuous nucleotides but are less than or equal to 300 kb, 200 kb, 100 kb, 50 kb, 15 kb, 10 kb, 7.5 kb, 5 kb, 2.5 kb, 2.0 kb, or 1 kb, in length. In a further embodiment, polynucleotides of the invention comprise a portion of the coding sequences, as disclosed herein, but do not comprise all or a portion of any intron. In another embodiment, the polynucleotides comprising coding sequences do not contain coding sequences of a genomic flanking gene (i.e., 5′ or 3′ to the gene of interest in the genome). In other embodiments, the polynucleotides of the invention do not contain the coding sequence of more than 1000, 500, 250, 100, 50, 25, 20, 15, 10, 5, 4, 3, 2, or 1 genomic flanking gene(s).

A “polynucleotide” of the present invention also includes those polynucleotides capable of hybridizing, under stringent hybridization conditions, to sequences contained in SEQ ID NO:X, or the complement thereof (e.g., the complement of any one, two, three, four, or more of the polynucleotide fragments described herein), the polynucleotide sequence delineated in columns 7 and 8 of Table 1A or the complement thereof, the polynucleotide sequence delineated in columns 8 and 9 of Table 2 or the complement thereof, and/or cDNA sequences contained in Clone ID: (e.g., the complement of any one, two, three, four, or more of the polynucleotide fragments, or the cDNA clone within the pool of cDNA clones deposited with the ATCC™, described herein), and/or the polynucleotide sequence delineated in column 6 of Table 1C or the complement thereof. “Stringent hybridization conditions” refers to an overnight incubation at 42 degree C. in a solution comprising 50% formamide, 5×SSC (750 mM NaCl, 75 mM trisodium citrate), 50 mM sodium phosphate (pH 7.6), 5×Denhardt's solution, 10% dextran sulfate, and 20 μg/ml denatured, sheared salmon sperm DNA, followed by washing the filters in 0.1×SSC at about 65 degree C.

Also contemplated are nucleic acid molecules that hybridize to the polynucleotides of the present invention at lower stringency hybridization conditions. Changes in the stringency of hybridization and signal detection are primarily accomplished through the manipulation of formamide concentration (lower percentages of formamide result in lowered stringency); salt conditions, or temperature. For example, lower stringency conditions include an overnight incubation at 37 degree C. in a solution comprising 6×SSPE (20×SSPE=3M NaCl; 0.2M NaH₂PO₄; 0.02M EDTA, pH 7.4), 0.5% SDS, 30% formamide, 100 μg/ml salmon sperm blocking DNA; followed by washes at 50 degree C. with 1×SSPE, 0.1% SDS. In addition, to achieve even lower stringency, washes performed following stringent hybridization can be done at higher salt concentrations (e.g. 5×SSC).

Note that variations in the above conditions may be accomplished through the inclusion and/or substitution of alternate blocking reagents used to suppress background in hybridization experiments. Typical blocking reagents include Denhardt's reagent, BLOTTO, heparin, denatured salmon sperm DNA, and commercially available proprietary formulations. The inclusion of specific blocking reagents may require modification of the hybridization conditions described above, due to problems with compatibility.

Of course, a polynucleotide which hybridizes only to polyA+ sequences (such as any 3′ terminal polyA+ tract of a cDNA shown in the sequence listing), or to a complementary stretch of T (or U) residues, would not be included in the definition of “polynucleotide,” since such a polynucleotide would hybridize to any nucleic acid molecule containing a poly (A) stretch or the complement thereof (e.g., practically any double-stranded cDNA clone generated using oligo dT as a primer).

The polynucleotide of the present invention can be composed of any polyribonucleotide or polydeoxyribonucleotide, which may be unmodified RNA or DNA or modified RNA or DNA. For example, polynucleotides can be composed of single- and double-stranded DNA, DNA that is a mixture of single- and double-stranded regions, single- and double-stranded RNA, and RNA that is mixture of single- and double-stranded regions, hybrid molecules comprising DNA and RNA that may be single-stranded or, more typically, double-stranded or a mixture of single- and double-stranded regions. In addition, the polynucleotide can be composed of triple-stranded regions comprising RNA or DNA or both RNA and DNA. A polynucleotide may also contain one or more modified bases or DNA or RNA backbones modified for stability or for other reasons. “Modified” bases include, for example, tritylated bases and unusual bases such as inosine. A variety of modifications can be made to DNA and RNA; thus, “polynucleotide” embraces chemically, enzymatically, or metabolically modified forms.

In specific embodiments, the polynucleotides of the invention are at least 15, at least 30, at least 50, at least 100, at least 125, at least 500, or at least 1000 continuous nucleotides but are less than or equal to 300 kb, 200 kb, 100 kb, 50 kb, 15 kb, 10 kb, 7.5 kb, 5 kb, 2.5 kb, 2.0 kb, or 1 kb, in length. In a further embodiment, polynucleotides of the invention comprise a portion of the coding sequences, as disclosed herein, but do not comprise all or a portion of any intron. In another embodiment, the polynucleotides comprising coding sequences do not contain coding sequences of a genomic flanking gene (i.e., 5′ or 3′ to the gene of interest in the genome). In other embodiments, the polynucleotides of the invention do not contain the coding sequence of more than 1000, 500, 250, 100, 50, 25, 20, 15, 10, 5, 4, 3, 2, or 1 genomic flanking gene(s).

“SEQ ID NO:X” refers to a polynucleotide sequence described in column 5 of Table 1A, while “SEQ ID NO:Y” refers to a polypeptide sequence described in column 10 of Table 1A. SEQ ID NO:X is identified by an integer specified in column 6 of Table 1A. The polypeptide sequence SEQ ID NO:Y is a translated open reading frame (ORF) encoded by polynucleotide SEQ ID NO:X. The polynucleotide sequences are shown in the sequence listing immediately followed by all of the polypeptide sequences. Thus, a polypeptide sequence corresponding to polynucleotide sequence SEQ ID NO:2 is the first polypeptide sequence shown in the sequence listing. The second polypeptide sequence corresponds to the polynucleotide sequence shown as SEQ ID NO:3, and so on.

The polypeptide of the present invention can be composed of amino acids joined to each other by peptide bonds or modified peptide bonds, i.e., peptide isosteres, and may contain amino acids other than the 20 gene-encoded amino acids. The polypeptides may be modified by either natural processes, such as posttranslational processing, or by chemical modification techniques which are well known in the art. Such modifications are well described in basic texts and in more detailed monographs, as well as in a voluminous research literature. Modifications can occur anywhere in a polypeptide, including the peptide backbone, the amino acid side-chains and the amino or carboxyl termini. It will be appreciated that the same type of modification may be present in the same or varying degrees at several sites in a given polypeptide. Also, a given polypeptide may contain many types of modifications. Polypeptides may be branched, for example, as a result of ubiquitination, and they may be cyclic, with or without branching. Cyclic, branched, and branched cyclic polypeptides may result from posttranslation natural processes or may be made by synthetic methods. Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cysteine, formation of pyroglutamate, formylation, gamma-carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, pegylation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination. (See, for instance, PROTEINS—STRUCTURE AND MOLECULAR PROPERTIES, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, New York (1993); POSTTRANSLATIONAL COVALENT MODIFICATION OF PROTEINS, B. C. Johnson, Ed., Academic Press, New York, pgs. 1-12 (1983); Seifter et al., Meth. Enzymol. 182:626-646 (1990); Rattan et al., Ann. N.Y. Acad. Sci. 663:48-62 (1992)).

“SEQ ID NO:X” refers to a polynucleotide sequence described, for example, in Tables 1A, Table 1B, or Table 2, while “SEQ ID NO:Y” refers to a polypeptide sequence described in column 11 of Table 1A and or Table 1B. SEQ ID NO:X is identified by an integer specified in column 4 of Table 1B. The polypeptide sequence SEQ ID NO:Y is a translated open reading frame (ORF) encoded by polynucleotide SEQ ID NO:X. “Clone ID:” refers to a cDNA clone described in column 2 of Table 1A and/or 1B.

“A polypeptide having functional activity” refers to a polypeptide capable of displaying one or more known functional activities associated with a full-length (complete) protein. Such functional activities include, but are not limited to, biological activity (e.g. activity useful in treating, preventing and/or ameliorating immune diseases and disorders), antigenicity (ability to bind [or compete with a polypeptide for binding] to an anti-polypeptide antibody), immunogenicity (ability to generate antibody which binds to a specific polypeptide of the invention), ability to form multimers with polypeptides of the invention, and ability to bind to a receptor or ligand for a polypeptide.

The polypeptides of the invention can be assayed for functional activity (e.g. biological activity) using or routinely modifying assays known in the art, as well as assays described herein. Specifically, one of skill in the art may routinely assay secreted polypeptides (including fragments and variants) of the invention for activity using assays as described in the examples section below.

“A polypeptide having biological activity” refers to a polypeptide exhibiting activity similar to, but not necessarily identical to, an activity of a polypeptide of the present invention, including mature forms, as measured in a particular biological assay, with or without dose dependency. In the case where dose dependency does exist, it need not be identical to that of the polypeptide, but rather substantially similar to the dose-dependence in a given activity as compared to the polypeptide of the present invention (i.e., the candidate polypeptide will exhibit greater activity or not more than about 25-fold less and, preferably, not more than about tenfold less activity, and most preferably, not more than about three-fold less activity relative to the polypeptide of the present invention).

Tables Table 1A

Table 1A summarizes information concerning certain polynucleotides and polypeptides of the invention. The first column provides the gene number in the application for each clone identifier. The second column provides a unique clone identifier, “Clone ID:”, for a cDNA clone related to each contig sequence disclosed in Table 1A. Third column, the cDNA Clones identified in the second column were deposited as indicated in the third column (i.e. by ATCC™ Deposit No:Z and deposit date). Some of the deposits contain multiple different clones corresponding to the same gene. In the fourth column, “Vector” refers to the type of vector contained in the corresponding cDNA Clone identified in the second column. In the fifth column, the nucleotide sequence identified as “NT SEQ ID NO:X” was assembled from partially homologous (“overlapping”) sequences obtained from the corresponding cDNA clone identified in the second column and, in some cases, from additional related cDNA clones. The overlapping sequences were assembled into a single contiguous sequence of high redundancy (usually three to five overlapping sequences at each nucleotide position), resulting in a final sequence identified as SEQ ID NO:X. In the sixth column, “Total NT Seq.” refers to the total number of nucleotides in the contig sequence identified as SEQ ID NO:X.” The deposited clone may contain all or most of these sequences, reflected by the nucleotide position indicated as “5′ NT of Clone Seq.” (seventh column) and the “3′ NT of Clone Seq.” (eighth column) of SEQ ID NO:X. In the ninth column, the nucleotide position of SEQ ID NO:X of the putative start codon (methionine) is identified as “5′ NT of Start Codon.” Similarly, in column ten, the nucleotide position of SEQ ID NO:X of the predicted signal sequence is identified as “5′ NT of First AA of Signal Pep.” In the eleventh column, the translated amino acid sequence, beginning with the methionine, is identified as “AA SEQ ID NO:Y,” although other reading frames can also be routinely translated using known molecular biology techniques. The polypeptides produced by these alternative open reading frames are specifically contemplated by the present invention.

In the twelfth and thirteenth columns of Table 1A, the first and last amino acid position of SEQ ID NO:Y of the predicted signal peptide is identified as “First AA of Sig Pep” and “Last AA of Sig Pep.” In the fourteenth column, the predicted first amino acid position of SEQ ID NO:Y of the secreted portion is identified as “Predicted First AA of Secreted Portion”. The amino acid position of SEQ ID NO:Y of the last amino acid encoded by the open reading frame is identified in the fifteenth column as “Last AA of ORF”.

SEQ ID NO:X (where X may be any of the polynucleotide sequences disclosed in the sequence listing) and the translated SEQ ID NO:Y (where Y may be any of the polypeptide sequences disclosed in the sequence listing) are sufficiently accurate and otherwise suitable for a variety of uses well known in the art and described further below. For instance, SEQ ID NO:X is useful for designing nucleic acid hybridization probes that will detect nucleic acid sequences contained in SEQ ID NO:X or the cDNA contained in the deposited clone. These probes will also hybridize to nucleic acid molecules in biological samples, thereby enabling a variety of forensic and diagnostic methods of the invention. Similarly, polypeptides identified from SEQ ID NO:Y may be used, for example, to generate antibodies which bind specifically to proteins containing the polypeptides and the secreted proteins encoded by the cDNA clones identified in Table 1A and/or elsewhere herein

Nevertheless, DNA sequences generated by sequencing reactions can contain sequencing errors. The errors exist as misidentified nucleotides, or as insertions or deletions of nucleotides in the generated DNA sequence. The erroneously inserted or deleted nucleotides cause frame shifts in the reading frames of the predicted amino acid sequence. In these cases, the predicted amino acid sequence diverges from the actual amino acid sequence, even though the generated DNA sequence may be greater than 99.9% identical to the actual DNA sequence (for example, one base insertion or deletion in an open reading frame of over 1000 bases).

Accordingly, for those applications requiring precision in the nucleotide sequence or the amino acid sequence, the present invention provides not only the generated nucleotide sequence identified as SEQ ID NO:X, and the predicted translated amino acid sequence identified as SEQ ID NO:Y, but also a sample of plasmid DNA containing a human cDNA of the invention deposited with the ATCC™, as set forth in Table 1A. The nucleotide sequence of each deposited plasmid can readily be determined by sequencing the deposited plasmid in accordance with known methods

The predicted amino acid sequence can then be verified from such deposits. Moreover, the amino acid sequence of the protein encoded by a particular plasmid can also be directly determined by peptide sequencing or by expressing the protein in a suitable host cell containing the deposited human cDNA, collecting the protein, and determining its sequence.

Also provided in Table 1A is the name of the vector which contains the cDNA plasmid. Each vector is routinely used in the art. The following additional information is provided for convenience.

Vectors Lambda Zap (U.S. Pat. Nos. 5,128,256 and 5,286,636), Uni-Zap XR (U.S. Pat. Nos. 5,128,256 and 5,286,636), Zap Express (U.S. Pat. Nos. 5,128,256 and 5,286,636), pBLUESCRIPT™ (pBS) (Short, J. M. et al., Nucleic Acids Res. 16:7583-7600 (1988); Alting-Mees, M. A. and Short, J. M., Nucleic Acids Res. 17:9494 (1989)) and pBK (Alting-Mees, M. A. et al., Strategies 5:58-61 (1992)) are commercially available from STRATAGENE™ Cloning Systems, Inc., 11011 N. Torrey Pines Road, La Jolla, Calif., 92037. pBS contains an ampicillin resistance gene and pBK contains a neomycin resistance gene. Phagemid pBS may be excised from the Lambda Zap and Uni-Zap XR vectors, and phagemid pBK may be excised from the Zap Express vector. Both phagemids may be transformed into E. coli strain XL-1 Blue, also available from STRATAGENET™.

Vectors pSport1, pCMVSport 1.0, pCMVSport 2.0 and pCMVSport 3.0, were obtained from Life Technologies, Inc., P.O. Box 6009, Gaithersburg, Md. 20897. All Sport vectors contain an ampicillin resistance gene and may be transformed into E. coli strain DH10B, also available from Life Technologies. See, for instance, Gruber, C. E., et al., Focus 15:59 (1993). Vector lafmid BA (Bento Soares, Columbia University, New York, N.Y.) contains an ampicillin resistance gene and can be transformed into E. coli strain XL-1 Blue. Vector pCR®2.1, which is available from Invitrogen, 1600 Faraday Avenue, Carlsbad, Calif. 92008, contains an ampicillin resistance gene and may be transformed into E. coli strain DH10B, available from Life Technologies. See, for instance, Clark, J. M., Nuc. Acids Res. 16:9677-9686 (1988) and Mead, D. et al., Bio/Technology 9: (1991).

The present invention also relates to the genes corresponding to SEQ ID NO:X, SEQ ID NO:Y, and/or a deposited cDNA (cDNA Clone ID). The corresponding gene can be isolated in accordance with known methods using the sequence information disclosed herein. Such methods include, but are not limited to, preparing probes or primers from the disclosed sequence and identifying or amplifying the corresponding gene from appropriate sources of genomic material.

Also provided in the present invention are allelic variants, orthologs, and/or species homologs. Procedures known in the art can be used to obtain full-length genes, allelic variants, splice variants, full-length coding portions, orthologs, and/or species homologs of genes corresponding to SEQ ID NO:X and SEQ ID NO:Y using information from the sequences disclosed herein or the clones deposited with the ATCC™. For example, allelic variants and/or species homologs may be isolated and identified by making suitable probes or primers from the sequences provided herein and screening a suitable nucleic acid source for allelic variants and/or the desired homologue.

The present invention provides a polynucleotide comprising, or alternatively consisting of, the nucleic acid sequence of SEQ ID NO:X and/or a cDNA contained in ATCC™ Deposit No.Z. The present invention also provides a polypeptide comprising, or alternatively, consisting of, the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X, and/or a polypeptide encoded by a cDNA contained in ATCC™ deposit No.Z. Polynucleotides encoding a polypeptide comprising, or alternatively consisting of the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X and/or a polypeptide encoded by the cDNA contained in ATCC™ Deposit No.Z, are also encompassed by the invention. The present invention further encompasses a polynucleotide comprising, or alternatively consisting of the complement of the nucleic acid sequence of SEQ ID NO:X, and/or the complement of the coding strand of the cDNA contained in ATCC™ Deposit No.Z.

TABLE 1A 5′ NT First Last First ATCC ™ NT 5′ NT 3′ NT of First AA AA AA AA of Last Deposit SEQ Total of of 5′ NT AA of SEQ of of Se- AA Gene cDNA No: Z and ID NT Clone Clone of Start Signal ID Sig Sig creted of No. Clone ID Date Vector NO: X Seq. Seq. Seq. Codon Pep NO: Y Pep Pep Portion ORF 1 H2CBG48 209889 pBLUESCRIPT ™ 11 2797 1 2797 125 125 908 1 25 26 45 May 22, 1998 SK− 2 H2MAC30 209299 pBLUESCRIPT ™ 12 459 1 459 157 157 909 1 28 29 72 Sep. 25, 1997 SK− 3 H6EAB28 209511 Uni-ZAP XR 13 1939 1 1939 115 115 910 1 31 32 100 Dec. 03, 1997 3 H6EAB28 209511 Uni-ZAP XR 603 1547 1 1547 116 116 1500 1 20 21 76 Dec. 03, 1997 4 H6EDF66 209299 Uni-ZAP XR 14 540 1 540 146 146 911 1 27 28 131 Sep. 25, 1997 5 H6EDX46 209626 Uni-ZAP XR 15 888 1 888 229 229 912 1 20 21 182 Feb. 12, 1998 5 H6EDX46 209626 Uni-ZAP XR 604 718 1 718 128 128 1501 1 20 21 84 Feb. 12, 1998 6 HABAG37 209626 pSport1 16 654 1 639 97 97 913 1 31 32 62 Feb. 12, 1998 7 HACBD91 209626 Uni-ZAP XR 17 1445 1 1445 117 117 914 1 42 43 49 Feb. 12, 1998 8 HACCI17 203071 Uni-ZAP XR 18 1722 336 1714 461 461 915 1 24 25 218 Jul. 27, 1998 8 HACCI17 203071 Uni-ZAP XR 605 1380 12 1380 135 135 1502 1 24 25 72 Jul. 27, 1998 9 HADAO89 209423 pSport1 19 1453 1 1453 244 244 916 1 22 23 44 Oct. 30, 1997 10 HADCP14 203027 pSport1 20 1032 1 1032 35 35 917 1 20 21 142 Jun. 26, 1998 11 HAGAI85 97922 Uni-ZAP XR 21 1752 52 1752 166 166 918 1 23 24 30 Mar. 07, 1997 209070 May 22, 1997 12 HAGAM64 209603 Uni-ZAP XR 22 2321 1 2321 57 57 919 1 31 32 44 Jan. 29, 1998 13 HAGAN21 PTA-841 Uni-ZAP XR 23 843 1 843 34 34 920 1 17 18 91 Oct. 13, 1999 13 HAGAN21 PTA-841 Uni-ZAP XR 606 610 294 610 335 335 1503 1 17 18 91 Oct. 13, 1999 13 HAGAN21 PTA-841 Uni-ZAP XR 607 659 1 659 452 1504 1 4 Oct. 13, 1999 13 HAGAN21 PTA-841 Uni-ZAP XR 608 189 1 189 146 1505 1 13 14 14 Oct. 13, 1999 13 HAGAN21 PTA-841 Uni-ZAP XR 609 637 1 637 321 1506 1 6 Oct. 13, 1999 14 HAGBZ81 209118 Uni-ZAP XR 24 1382 24 1382 65 921 1 30 31 49 Jun. 12, 1997 15 HAGDG59 209277 Uni-ZAP XR 25 1734 44 1717 124 124 922 1 18 19 300 Sep. 18, 1997 16 HAGDS20 209299 Uni-ZAP XR 26 919 1 919 11 11 923 1 17 18 66 Sep. 25, 1997 17 HAGFG51 203364 Uni-ZAP XR 27 1313 1 1313 163 163 924 1 23 24 43 Oct. 19, 1998 18 HAHDB16 209626 Uni-ZAP XR 28 796 1 796 93 93 925 1 20 21 50 Feb. 12, 1998 19 HAHDR32 209626 Uni-ZAP XR 29 1256 365 1256 435 435 926 1 25 26 181 Feb. 12, 1998 20 HAIBO71 209145 Uni-ZAP XR 30 752 172 752 325 325 927 1 28 29 66 Jul. 17, 1997 21 HAIBP89 209877 Uni-ZAP XR 31 2243 173 2243 311 311 928 1 27 28 317 May 18, 1998 21 HAIBP89 209877 Uni-ZAP XR 610 1025 1 1025 1 1507 1 1 2 18 May 18, 1998 22 HAICP19 209009 Uni-ZAP XR 32 1624 89 1483 128 128 929 1 18 19 446 Apr. 28, 1997 23 HAIFL18 209852 Uni-ZAP XR 33 879 1 879 274 274 930 1 29 30 140 May 07, 1998 24 HAJAF57 203364 pCMVSport 3.0 34 2761 1 2761 43 43 931 1 1 2 94 Oct. 19, 1998 25 HAJBR69 209626 pCMVSport 3.0 35 755 1 755 262 262 932 1 19 20 53 Feb. 12, 1998 26 HAJBZ75 209603 pCMVSport 3.0 36 2089 10 2085 49 49 933 1 22 23 607 Jan. 29, 1998 27 HAMFC93 PTA-849 pCMVSport 3.0 37 2534 1 2534 136 136 934 1 30 31 191 Oct. 13, 1999 27 HAMFC93 PTA-849 pCMVSport 3.0 611 824 1 824 115 115 1508 1 30 31 178 Oct. 13, 1999 27 HAMFC93 PTA-849 pCMVSport 3.0 612 3941 1947 3941 323 1509 1 8 Oct. 13, 1999 28 HAMFK58 209641 pCMVSport 3.0 38 785 1 785 279 279 935 1 31 32 79 Feb. 25, 1998 29 HAPNY86 209511 Uni-ZAP XR 39 1280 1 1280 100 100 936 1 25 26 129 Dec. 03, 1997 30 HAPPW30 209683 Uni-ZAP XR 40 1472 1 1472 59 59 937 1 22 23 264 Mar. 20, 1998 30 HAPPW30 209683 Uni-ZAP XR 613 1508 14 1501 54 54 1510 1 22 23 91 Mar. 20, 1998 31 HAPQT22 203070 Uni-ZAP XR 41 635 1 635 132 132 938 1 17 18 72 Jul. 27, 1998 32 HASAV70 97923 Uni-ZAP XR 42 729 1 729 94 94 939 1 20 21 110 Mar. 07, 1997 209071 May 22, 1997 32 HASAV70 97923 Uni-ZAP XR 614 1412 10 733 103 103 1511 1 20 21 110 Mar. 07, 1997 209071 May 22, 1997 33 HASCG84 209568 Uni-ZAP XR 43 1079 1 1079 216 216 940 1 32 33 53 Jan. 06, 1998 34 HATAC53 209651 Uni-ZAP XR 44 1959 1 1959 97 97 941 1 21 22 248 Mar. 04, 1998 34 HATAC53 209651 Uni-ZAP XR 615 1306 13 1306 99 99 1512 1 21 22 189 Mar. 04, 1998 35 HATBR65 209626 Uni-ZAP XR 45 812 1 812 252 252 942 1 16 17 64 Feb. 12, 1998 36 HATCB92 209683 Uni-ZAP XR 46 1756 1 1756 247 247 943 1 37 38 56 Mar. 20, 1998 37 HATCP77 209965 Uni-ZAP XR 47 2098 1 2098 37 37 944 1 21 22 182 Jun. 11, 1998 38 HATDF29 203858 Uni-ZAP XR 48 1355 1 1355 143 143 945 1 30 31 385 Mar. 18, 1999 39 HATDM46 PTA-844 Uni-ZAP XR 49 2325 1 2325 130 130 946 1 18 19 68 Oct. 13, 1999 39 HATDM46 PTA-844 Uni-ZAP XR 616 546 1 546 131 131 1513 1 18 19 68 Oct. 13, 1999 39 HATDM46 PTA-844 Uni-ZAP XR 617 859 1 859 723 1514 1 10 11 29 Oct. 13, 1999 39 HATDM46 PTA-844 Uni-ZAP XR 618 1649 1 1649 988 1515 1 15 16 62 Oct. 13, 1999 39 HATDM46 PTA-844 Uni-ZAP XR 619 675 204 651 1 1516 1 1 2 225 Oct. 13, 1999 39 HATDM46 PTA-844 Uni-ZAP XR 620 751 563 751 2 1517 1 1 2 211 Oct. 13, 1999 40 HATEE46 209407 Uni-ZAP XR 50 1675 136 863 241 241 947 1 21 22 53 Oct. 23, 1997 41 HBAFJ33 209603 pSport1 51 1280 1 1252 60 60 948 1 15 16 110 Jan. 29, 1998 42 HBAFV19 PTA-1543 pSport1 52 953 1 953 6 6 949 1 1 2 258 Mar. 21, 2000 43 HBAMB34 209324 pSport1 53 1027 1 1027 87 87 950 1 35 36 48 Oct. 02, 1997 44 HBCPB32 PTA-2075 pSport1 54 1368 1 1368 88 88 951 1 37 38 202 Jun. 09, 2000 44 HBCPB32 PTA-2075 pSport1 621 729 1 729 89 89 1518 1 37 38 196 Jun. 09, 2000 45 HBHAD12 209009 Uni-ZAP XR 55 786 1 786 176 952 1 17 18 23 Apr. 28, 1997 46 HBHMA23 209782 pSport1 56 1175 2 1175 71 71 953 1 24 25 197 Apr. 20, 1998 46 HBHMA23 209782 pSport1 622 1172 1 1172 70 70 1519 1 24 25 76 Apr. 20, 1998 47 HBIBW67 209324 Uni-ZAP XR 57 1404 1 1404 685 685 954 1 33 34 38 Oct. 02, 1997 48 HBIMB51 209683 pCMVSport 3.0 58 537 1 537 98 98 955 1 21 22 146 Mar. 20, 1998 48 HBIMB51 209683 pCMVSport 3.0 623 526 1 526 93 93 1520 1 21 22 130 Mar. 20, 1998 49 HBINS58 PTA-885 pCMVSport 3.0 59 843 1 843 57 57 956 1 30 31 174 Oct. 28, 1999 49 HBINS58 PTA-885 pCMVSport 3.0 624 1566 1 1566 71 71 1521 1 29 30 173 Oct. 28, 1999 49 HBINS58 PTA-885 pCMVSport 3.0 625 1067 1 1067 100 100 1522 1 29 30 210 Oct. 28, 1999 50 HBJFU48 209125 Uni-ZAP XR 60 849 1 849 20 20 957 1 39 40 40 Jun. 19, 1997 51 HBJID05 209300 Uni-ZAP XR 61 2008 1 2008 157 157 958 1 20 21 199 Sep. 25, 1997 51 HBJID05 209300 Uni-ZAP XR 626 571 1 571 137 137 1523 1 20 21 111 Sep. 25, 1997 52 HBJIY92 203071 Uni-ZAP XR 62 2434 487 2366 548 548 959 1 29 30 40 Jul. 27, 1998 53 HBJJU28 209346 Uni-ZAP XR 63 1160 1 1160 133 133 960 1 18 19 84 Oct. 09, 1997 54 HBJLC01 209651 Uni-ZAP XR 64 872 1 872 87 87 961 1 34 35 46 Mar. 04, 1998 55 HBJLF01 209877 Uni-ZAP XR 65 1932 201 1931 217 217 962 1 46 47 244 May 18, 1998 56 HBJLH40 203499 Uni-ZAP XR 66 1853 1 1853 74 74 963 1 30 31 74 Dec. 01, 1998 57 HBJNC59 PTA-622 Uni-ZAP XR 67 1061 1 1061 66 66 964 1 22 23 245 Sep. 02, 1999 57 HBJNC59 PTA-622 Uni-ZAP XR 627 1021 1 1021 66 66 1524 1 22 23 99 Sep. 02, 1999 57 HBJNC59 PTA-622 Uni-ZAP XR 628 1086 1 1023 64 64 1525 1 22 23 245 Sep. 02, 1999 58 HBNAW17 209242 Uni-ZAP XR 68 601 1 601 77 77 965 1 37 38 61 Sep. 12, 1997 59 HBOEG11 PTA-2072 pSport1 69 1356 1 1356 57 57 966 1 22 23 250 Jun. 09, 2000 59 HBOEG11 PTA-2072 pSport1 629 1352 1 1352 53 53 1526 1 22 23 250 Jun. 09, 2000 59 HBOEG11 PTA-2072 pSport1 630 1337 1 1289 47 47 1527 1 22 23 250 Jun. 09, 2000 60 HBOEG69 203081 pSport1 70 1411 1 1411 302 302 967 1 19 20 54 Jul. 30, 1998 61 HBXFL29 203858 ZAP Express 71 2229 376 2210 560 560 968 1 31 32 57 Mar. 18, 1999 62 HCACU58 209626 Uni-ZAP XR 72 1554 1 1554 137 137 969 1 30 31 83 Feb. 12, 1998 63 HCACV51 209551 Uni-ZAP XR 73 2083 1 2083 168 168 970 1 31 32 81 Dec. 12, 1997 63 HCACV51 209551 Uni-ZAP XR 631 2092 1 2092 173 173 1528 1 31 32 281 Dec. 12, 1997 64 HCDBW86 209242 Uni-ZAP XR 74 730 1 730 139 139 971 1 18 19 30 Sep. 12, 1997 65 HCE1Q89 209242 Uni-ZAP XR 75 863 1 863 74 74 972 1 17 18 88 Sep. 12, 1997 66 HCE2F54 209626 Uni-ZAP XR 76 1276 19 1256 166 166 973 1 19 20 319 Feb. 12, 1998 67 HCE3G69 209878 Uni-ZAP XR 77 2084 1 2084 165 165 974 1 19 20 336 May 18, 1998 67 HCE3G69 209878 Uni-ZAP XR 632 2078 1 2078 165 165 1529 1 19 20 105 May 18, 1998 68 HCEEA88 209626 Uni-ZAP XR 78 1016 1 1016 134 134 975 1 23 24 60 Feb. 12, 1998 69 HCEFB69 209965 Uni-ZAP XR 79 1430 1 1430 188 188 976 1 24 25 224 Jun. 11, 1998 70 HCEFB80 PTA-2069 Uni-ZAP XR 80 2494 1 2494 12 12 977 1 35 36 89 Jun. 09, 2000 70 HCEFB80 PTA-2069 Uni-ZAP XR 633 2494 1 2451 5 5 1530 1 35 36 89 Jun. 09, 2000 71 HCEGR33 209090 Uni-ZAP XR 81 1630 1 1630 243 243 978 1 18 19 31 Jun. 05, 1997 72 HCEMP62 209745 Uni-ZAP XR 82 1860 269 1726 352 352 979 1 30 31 187 Apr. 07, 1998 72 HCEMP62 209745 Uni-ZAP XR 634 1957 582 1823 19 19 1531 1 33 34 335 Apr. 07, 1998 73 HCENK38 209651 Uni-ZAP XR 83 1509 1 1509 10 10 980 1 28 29 52 Mar. 04, 1998 74 HCEWE17 PTA-842 Uni-ZAP XR 84 967 1 967 117 117 981 1 23 24 106 Oct. 13, 1999 74 HCEWE17 PTA-842 Uni-ZAP XR 635 730 247 730 500 500 1532 1 19 20 27 Oct. 13, 1999 74 HCEWE17 PTA-842 Uni-ZAP XR 636 550 1 550 156 1533 1 1 2 54 Oct. 13, 1999 75 HCEWE20 209300 Uni-ZAP XR 85 885 13 885 166 166 982 1 18 19 51 Sep. 25, 1997 76 HCFCU88 209324 pSport1 86 853 1 853 217 217 983 1 18 19 97 Oct. 02, 1997 77 HCFMV71 209242 pSport1 87 400 1 400 31 31 984 1 24 25 58 Sep. 12, 1997 78 HCFNN01 209086 pSport1 88 1261 154 1261 254 254 985 1 27 28 43 May 29, 1997 79 HCFOM18 209324 pSport1 89 639 1 639 28 28 986 1 20 21 63 Oct. 02, 1997 80 HCHNF25 209651 pSport1 90 3576 1 3576 1130 1130 987 1 30 31 169 Mar. 04, 1998 80 HCHNF25 209651 pSport1 637 807 1 807 180 180 1534 1 30 31 147 Mar. 04, 1998 81 HCMSQ56 209877 Uni-ZAP XR 91 1262 1 1262 148 148 988 1 19 20 88 May 18, 1998 82 HCMST14 209346 Uni-ZAP XR 92 614 1 614 136 136 989 1 24 25 47 Oct. 09, 1997 83 HCMTB45 209368 Uni-ZAP XR 93 958 1 958 215 215 990 1 20 21 123 Oct. 16, 1997 83 HCMTB45 209368 Uni-ZAP XR 638 946 1 946 209 209 1535 1 27 28 70 Oct. 16, 1997 84 HCNSD93 209627 pBLUESCRIPT ™ 94 1106 1 1106 139 139 991 1 21 22 46 Feb. 12, 1998 85 HCOOS80 PTA-2076 pSport1 95 1254 1 1254 36 36 992 1 26 27 158 Jun. 09, 2000 85 HCOOS80 PTA-2076 pSport1 639 869 15 869 40 40 1536 1 26 27 158 Jun. 09, 2000 85 HCOOS80 PTA-2076 pSport1 640 692 339 506 1 1537 1 1 2 106 Jun. 09, 2000 86 HCQCT05 PTA-884 Lambda ZAP II 96 679 1 679 381 993 1 2 Oct. 28, 1999 86 HCQCT05 PTA-884 Lambda ZAP II 641 2333 1324 2333 1702 1538 1 2 Oct. 28, 1999 87 HCUBS50 209215 ZAP Express 97 865 1 865 88 88 994 1 34 35 38 Aug. 21, 1997 88 HCUCK44 209853 ZAP Express 98 1139 573 1133 593 593 995 1 30 31 60 May 07, 1998 89 HCUEO60 209215 ZAP Express 99 1222 1 1222 102 102 996 1 34 35 64 Aug. 21, 1997 90 HCUGM86 PTA-1544 ZAP Express 100 627 1 627 91 91 997 1 24 25 44 Mar. 21, 2000 91 HCUHK65 209641 ZAP Express 101 367 1 367 80 80 998 1 26 27 79 Feb. 25, 1998 91 HCUHK65 209641 ZAP Express 642 3113 2577 2946 770 770 1539 1 30 31 708 Feb. 25, 1998 92 HCUIM65 209324 ZAP Express 102 875 331 736 557 557 999 1 27 28 47 Oct. 02, 1997 93 HCWEB58 PTA-883 ZAP Express 103 1283 1 1283 148 148 1000 1 27 28 343 Oct. 28, 1999 93 HCWEB58 PTA-883 ZAP Express 643 980 1 980 247 247 1540 1 27 28 244 Oct. 28, 1999 93 HCWEB58 PTA-883 ZAP Express 644 888 1 888 155 155 1541 1 27 28 244 Oct. 28, 1999 94 HCWGU37 PTA-883 ZAP Express 104 2777 1 2777 194 194 1001 1 10 Oct. 28, 1999 94 HCWGU37 PTA-883 ZAP Express 645 1651 1 1651 187 187 1542 1 10 Oct. 28, 1999 95 HCWKC15 209324 ZAP Express 105 710 1 710 37 37 1002 1 18 19 40 Oct. 02, 1997 96 HCWLD74 209626 ZAP Express 106 1540 1 1540 138 138 1003 1 21 22 65 Feb. 12, 1998 97 HDHEB60 209215 pCMVSport 2.0 107 1421 235 1421 568 568 1004 1 24 25 108 Aug. 21, 1997 98 HDHIA94 209627 pCMVSport 2.0 108 1489 1 1489 154 154 1005 1 30 31 168 Feb. 12, 1998 98 HDHIA94 209627 pCMVSport 2.0 646 2492 1 2492 163 163 1543 1 30 31 48 Feb. 12, 1998 99 HDHMA45 203331 pCMVSport 2.0 109 2184 1 2184 199 199 1006 1 33 34 413 Oct. 08, 1998 99 HDHMA45 203331 pCMVSport 2.0 647 2190 1 2190 204 204 1544 1 33 34 413 Oct. 08, 1998 100 HDHMA72 209324 pCMVSport 2.0 110 4463 216 2158 287 287 1007 1 36 37 315 Oct. 02, 1997 101 HDLAC10 209745 pCMVSport 2.0 111 1477 1 1477 132 132 1008 1 29 30 81 Apr. 07, 1998 102 HDLAO28 PTA-499 pCMVSport 2.0 112 1984 1 1984 259 259 1009 1 21 22 76 Aug. 11, 1999 103 HDPBA28 PTA-163 pCMVSport 3.0 113 3447 197 3447 259 259 1010 1 32 33 941 Jun. 01, 1999 103 HDPBA28 PTA-163 pCMVSport 3.0 648 4909 1 4909 69 69 1545 1 32 33 941 Jun. 01, 1999 104 HDPBQ02 209889 pCMVSport 3.0 114 1166 1 1166 461 461 1011 1 24 25 222 May 22, 1998 104 HDPBQ02 209889 pCMVSport 3.0 649 2191 291 2191 460 460 1546 1 24 25 108 May 22, 1998 105 HDPBQ71 209877 pCMVSport 3.0 115 2312 1 2312 93 93 1012 1 33 34 612 May 18, 1998 105 HDPBQ71 209877 pCMVSport 3.0 650 2242 6 2242 24 24 1547 1 33 34 612 May 18, 1998 105 HDPBQ71 209877 pCMVSport 3.0 651 2381 146 2381 165 165 1548 1 33 34 456 May 18, 1998 106 HDPCO25 209125 pCMVSport 3.0 116 767 76 767 182 182 1013 1 20 21 53 Jun. 19, 1997 107 HDPCY37 209568 pCMVSport 3.0 117 1932 45 1932 76 76 1014 1 21 22 578 Jan. 06, 1998 107 HDPCY37 209568 pCMVSport 3.0 652 1931 45 1931 76 76 1549 1 21 22 264 Jan. 06, 1998 108 HDPFF39 209511 pCMVSport 3.0 118 1256 1 1256 175 175 1015 1 18 19 196 Dec. 03, 1997 109 HDPGK25 209877 pCMVSport 3.0 119 703 1 703 345 345 1016 1 33 34 119 May 18, 1998 110 HDPGP94 203364 pCMVSport 3.0 120 3881 1 3881 256 256 1017 1 18 19 74 Oct. 19, 1998 111 HDPHI51 209125 pCMVSport 3.0 121 728 1 728 245 245 1018 1 30 31 40 Jun. 19, 1997 112 HDPJF37 209852 pCMVSport 3.0 122 986 1 986 196 196 1019 1 23 24 57 May 07, 1998 113 HDPJM30 209563 pCMVSport 3.0 123 1635 308 1633 59 59 1020 1 59 60 525 Dec. 18, 1997 113 HDPJM30 209563 pCMVSport 3.0 653 1314 1 1313 259 259 1550 1 20 21 59 Dec. 18, 1997 114 HDPNC61 209627 pCMVSport 3.0 124 1410 1 1410 20 20 1021 1 22 23 94 Feb. 12, 1998 115 HDPND46 209627 pCMVSport 3.0 125 1727 1 1727 15 15 1022 1 22 23 484 Feb. 12, 1998 116 HDPOE32 PTA-622 pCMVSport 3.0 126 1353 1 1353 118 118 1023 1 34 35 151 Sep. 02, 1999 117 HDPOH06 209745 pCMVSport 3.0 127 2504 1 2504 252 252 1024 1 29 30 242 Apr. 07, 1998 118 HDPOZ56 209889 pCMVSport 3.0 128 1905 1 1905 91 91 1025 1 21 22 567 May 22, 1998 118 HDPOZ56 209889 pCMVSport 3.0 654 1867 415 1867 103 103 1551 1 21 22 566 May 22, 1998 118 HDPOZ56 209889 pCMVSport 3.0 655 1722 1 1722 59 59 1552 1 21 22 319 May 22, 1998 119 HDPPA04 PTA-867 pCMVSport 3.0 129 2406 1 2406 271 271 1026 1 19 20 283 Oct. 26, 1999 119 HDPPA04 PTA-867 pCMVSport 3.0 656 1675 1 1613 1003 1553 1 11 12 23 Oct. 26, 1999 119 HDPPA04 PTA-867 pCMVSport 3.0 657 786 1 786 261 261 1554 1 19 20 93 Oct. 26, 1999 120 HDPPH47 209626 pCMVSport 3.0 130 2080 105 2080 116 116 1027 1 35 36 540 Feb. 12, 1998 121 HDPSB18 PTA-868 pCMVSport 3.0 131 3408 1 3408 123 123 1028 1 18 19 66 Oct. 26, 1999 121 HDPSB18 PTA-868 pCMVSport 3.0 658 308 1 308 116 1555 1 17 18 64 Oct. 26, 1999 121 HDPSB18 PTA-868 pCMVSport 3.0 659 1568 1 1568 1525 1556 1 7 8 14 Oct. 26, 1999 121 HDPSB18 PTA-868 pCMVSport 3.0 660 865 1 865 345 1557 1 1 2 107 Oct. 26, 1999 122 HDPSP01 209745 pCMVSport 3.0 132 2343 1 2343 184 184 1029 1 20 21 710 Apr. 07, 1998 122 HDPSP01 209745 pCMVSport 3.0 661 1752 1 1752 227 227 1558 1 20 21 308 Apr. 07, 1998 123 HDPSP54 209782 pCMVSport 3.0 133 3091 2304 3091 2356 2356 1030 1 18 19 48 Apr. 20, 1998 123 HDPSP54 209782 pCMVSport 3.0 662 536 1 536 179 179 1559 1 41 42 55 Apr. 20, 1998 124 HDPSU13 209627 pCMVSport 3.0 134 1218 1 1218 14 14 1031 1 25 26 114 Feb. 12, 1998 125 HDPTD15 209782 pCMVSport 3.0 135 1396 1 1396 223 223 1032 1 18 19 200 Apr. 20, 1998 126 HDPTK41 209965 pCMVSport 3.0 136 1564 1 1564 39 39 1033 1 26 27 369 Jun. 11, 1998 127 HDPUG50 209745 pCMVSport 3.0 137 1734 1 1734 22 22 1034 1 34 35 526 Apr. 07, 1998 128 HDPUH26 PTA-163 pCMVSport 3.0 138 2916 1 2916 90 90 1035 1 18 19 549 Jun. 01, 1999 129 HDPUW68 203331 pCMVSport 3.0 139 1748 1 1748 40 40 1036 1 18 19 467 Oct. 08, 1998 130 HDPVH60 203105 pCMVSport 3.0 140 3116 1 3100 8 8 1037 1 45 46 51 Aug. 13, 1998 131 HDPWN93 PTA-868 pCMVSport 3.0 141 2679 1 2669 45 45 1038 1 19 20 802 Oct. 26, 1999 131 HDPWN93 PTA-868 pCMVSport 3.0 663 716 1 716 35 35 1560 1 19 20 214 Oct. 26, 1999 131 HDPWN93 PTA-868 pCMVSport 3.0 664 2716 26 2716 27 27 1561 1 19 20 43 Oct. 26, 1999 132 HDQHD03 203570 pCMVSport 3.0 142 1266 1 1266 274 274 1039 1 20 21 331 Jan. 11, 1999 132 HDQHD03 203570 pCMVSport 3.0 665 1257 1 1257 259 259 1562 1 20 21 333 Jan. 11, 1999 133 HDTBP04 209300 pCMVSport 2.0 143 961 1 961 70 70 1040 1 15 16 219 Sep. 25, 1997 133 HDTBP04 209300 pCMVSport 2.0 666 959 1 959 65 65 1563 1 15 16 220 Sep. 25, 1997 134 HDTEK44 PTA-867 pCMVSport 2.0 144 2070 20 2070 691 1041 1 12 13 83 Oct. 26, 1999 134 HDTEK44 PTA-867 pCMVSport 2.0 667 1005 1 1005 175 175 1564 1 17 18 67 Oct. 26, 1999 134 HDTEK44 PTA-867 pCMVSport 2.0 668 2988 1 2988 116 116 1565 1 17 18 67 Oct. 26, 1999 134 HDTEK44 PTA-867 pCMVSport 2.0 669 2052 2 2052 673 1566 1 12 13 83 Oct. 26, 1999 135 HDTEN81 209463 pCMVSport 2.0 145 566 1 566 114 114 1042 1 17 18 85 Nov. 14, 1997 136 HDTFE17 PTA-868 pCMVSport 2.0 146 1242 1 1242 260 260 1043 1 20 21 29 Oct. 26, 1999 136 HDTFE17 PTA-868 pCMVSport 2.0 670 628 1 628 251 251 1567 1 20 21 29 Oct. 26, 1999 136 HDTFE17 PTA-868 pCMVSport 2.0 671 923 29 903 101 1568 1 6 7 80 Oct. 26, 1999 137 HDTGC73 209627 pCMVSport 2.0 147 712 1 712 386 386 1044 1 31 32 49 Feb. 12, 1998 138 HDTIT10 203570 pCMVSport 2.0 148 1200 1 813 58 58 1045 1 56 57 297 Jan. 11, 1999 138 HDTIT10 203570 pCMVSport 2.0 672 1159 1 805 161 161 1569 1 30 31 56 Jan. 11, 1999 139 HDTMK50 PTA-884 pCMVSport 2.0 149 1352 1 1352 154 154 1046 1 21 22 51 Oct. 28, 1999 139 HDTMK50 PTA-884 pCMVSport 2.0 673 912 1 912 164 164 1570 1 21 22 51 Oct. 28, 1999 139 HDTMK50 PTA-884 pCMVSport 2.0 674 321 1 321 200 1571 1 1 Oct. 28, 1999 140 HE2DY70 209877 Uni-ZAP XR 150 639 1 639 137 137 1047 1 45 46 58 May 18, 1998 141 HE2EN04 209300 Uni-ZAP XR 151 370 1 370 57 57 1048 1 16 17 50 Sep. 25, 1997 142 HE2FV03 97955 Uni-ZAP XR 152 2067 1 1251 116 116 1049 1 21 22 42 Mar. 13, 1997 209074 May 22, 1997 143 HE2NV57 209877 Uni-ZAP XR 153 867 1 867 99 99 1050 1 36 37 99 May 18, 1998 144 HE2PD49 209627 Uni-ZAP XR 154 1422 257 1404 337 337 1051 1 18 19 171 Feb. 12, 1998 145 HE2PY40 209965 Uni-ZAP XR 155 1288 1 1288 147 147 1052 1 22 23 83 Jun. 11, 1998 146 HE6EU50 97975 Uni-ZAP XR 156 1152 117 686 237 237 1053 1 20 21 34 Apr. 04, 1997 209081 May 29, 1997 147 HE8MH91 209603 Uni-ZAP XR 157 1761 1 1761 63 63 1054 1 23 24 116 Jan. 29, 1998 148 HE8QV67 PTA-2072 Uni-ZAP XR 158 1999 643 1999 502 502 1055 1 49 50 80 Jun. 09, 2000 148 HE8QV67 PTA-2072 Uni-ZAP XR 675 2342 1956 2276 256 1572 1 1 2 415 Jun. 09, 2000 149 HE8UB86 203570 Uni-ZAP XR 159 1021 1 1021 201 201 1056 1 21 22 250 Jan. 11, 1999 150 HE9BK23 209683 Uni-ZAP XR 160 1636 1 1636 39 39 1057 1 21 22 309 Mar. 20, 1998 151 HE9CO69 209551 Uni-ZAP XR 161 1077 1 1077 161 161 1058 1 26 27 41 Dec. 12, 1997 152 HE9CP41 209368 Uni-ZAP XR 162 1392 1 1392 132 132 1059 1 20 21 41 Oct. 16, 1997 153 HE9DG49 97923 Uni-ZAP XR 163 717 1 717 70 70 1060 1 28 29 201 Mar. 07, 1997 209071 May 22, 1997 153 HE9DG49 97923 Uni-ZAP XR 676 717 1 717 70 70 1573 1 27 28 201 Mar. 07, 1997 209071 May 22, 1997 153 HE9DG49 97923 Uni-ZAP XR 677 713 17 713 78 78 1574 1 28 29 203 Mar. 07, 1997 209071 May 22, 1997 154 HE9OW20 203570 Uni-ZAP XR 164 1209 1 1209 129 129 1061 1 33 34 355 Jan. 11, 1999 154 HE9OW20 203570 Uni-ZAP XR 678 1165 1 1165 136 136 1575 1 30 31 313 Jan. 11, 1999 154 HE9OW20 203570 Uni-ZAP XR 679 1160 1 1160 129 129 1576 1 30 31 134 Jan. 11, 1999 155 HE9RM63 PTA-499 Uni-ZAP XR 165 2149 1 2149 82 82 1062 1 27 28 354 Aug. 11, 1999 156 HEAAR07 209346 Uni-ZAP XR 166 1084 1 1084 48 48 1063 1 31 32 42 Oct. 09, 1997 157 HEBAE88 209242 Uni-ZAP XR 167 582 1 582 160 160 1064 1 26 27 42 Sep. 12, 1997 158 HEBBN36 209141 Uni-ZAP XR 168 1046 470 1046 645 645 1065 1 29 30 53 Jul. 09, 1997 159 HEBCM63 209141 Uni-ZAP XR 169 558 1 558 246 246 1066 1 26 27 68 Jul. 09, 1997 160 HEBEJ18 203069 Uni-ZAP XR 170 685 7 649 51 51 1067 1 15 16 139 Jul. 27, 1998 161 HEEAG23 209745 Uni-ZAP XR 171 1669 25 1280 57 57 1068 1 18 19 46 Apr. 07, 1998 162 HEEAJ02 209627 Uni-ZAP XR 172 1038 148 1037 387 387 1069 1 40 41 125 Feb. 12, 1998 163 HEEAQ11 203071 Uni-ZAP XR 173 921 1 921 213 213 1070 1 28 29 147 Jul. 27, 1998 164 HEGAN94 203071 Uni-ZAP XR 174 582 1 582 52 52 1071 1 23 24 121 Jul. 27, 1998 164 HEGAN94 203071 Uni-ZAP XR 680 680 1 680 133 133 1577 1 23 24 121 Jul. 27, 1998 165 HEGBS69 PTA-2082 Uni-ZAP XR 175 809 1 809 260 260 1072 1 20 21 161 Jun. 09, 2000 165 HEGBS69 PTA-2082 Uni-ZAP XR 681 1188 1 807 253 253 1578 1 20 21 161 Jun. 09, 2000 166 HELGK31 209878 Uni-ZAP XR 176 1396 25 1334 209 209 1073 1 29 30 344 May 18, 1998 166 HELGK31 209878 Uni-ZAP XR 682 1342 68 1342 402 402 1579 1 1 2 291 May 18, 1998 167 HELHD85 PTA-1544 Uni-ZAP XR 177 1886 1 1886 41 41 1074 1 25 26 79 Mar. 21, 2000 168 HELHL48 209877 Uni-ZAP XR 178 2971 560 2557 629 629 1075 1 16 17 291 May 18, 1998 168 HELHL48 209877 Uni-ZAP XR 683 1955 1 1955 31 31 1580 1 16 17 184 May 18, 1998 169 HEMAM41 209010 Uni-ZAP XR 179 1337 60 1328 175 175 1076 1 39 40 190 Apr. 28, 1997 209085 May 29, 1997 169 HEMAM41 209010 Uni-ZAP XR 684 1338 33 1327 175 175 1581 1 32 33 91 Apr. 28, 1997 209085 May 29, 1997 170 HEPAA46 209551 Uni-ZAP XR 180 1129 1 1129 18 18 1077 1 20 21 123 Dec. 12, 1997 171 HEQAK71 209551 pCMVSport 3.0 181 1689 1 1689 198 198 1078 1 17 18 44 Dec. 12, 1997 172 HEQCC55 209965 pCMVSport 3.0 182 1000 1 1000 25 25 1079 1 27 28 129 Jun. 11, 1998 172 HEQCC55 209965 pCMVSport 3.0 685 1052 30 1052 62 62 1582 1 27 28 112 Jun. 11, 1998 172 HEQCC55 209965 pCMVSport 3.0 686 1037 1 1037 57 57 1583 1 27 28 155 Jun. 11, 1998 173 HERAD40 209853 Uni-ZAP XR 183 990 1 990 85 85 1080 1 38 39 98 May 07, 1998 174 HERAR44 209407 Uni-ZAP XR 184 420 1 420 60 60 1081 1 40 41 45 Oct. 23, 1997 175 HESAJ10 209242 Uni-ZAP XR 185 1090 400 1090 405 405 1082 1 23 24 71 Sep. 12, 1997 176 HETAB45 209580 Uni-ZAP XR 186 1676 1 1676 123 123 1083 1 30 31 179 Jan. 14, 1998 177 HETBR16 209877 Uni-ZAP XR 187 1569 1 1569 161 161 1084 1 21 22 64 May 18, 1998 178 HETEU28 PTA-842 Uni-ZAP XR 188 1381 1 1381 256 256 1085 1 34 35 153 Oct. 13, 1999 178 HETEU28 PTA-842 Uni-ZAP XR 687 1501 1 1462 331 331 1584 1 34 35 153 Oct. 13, 1999 179 HETLM70 PTA-2073 Uni-ZAP XR 189 1251 1 1199 336 336 1086 1 27 28 229 Jun. 09, 2000 179 HETLM70 PTA-2073 Uni-ZAP XR 688 1251 1 1251 336 336 1585 1 27 28 229 Jun. 09, 2000 179 HETLM70 PTA-2073 Uni-ZAP XR 689 517 161 517 2 1586 1 1 2 85 Jun. 09, 2000 180 HFABG18 PTA-1544 Uni-ZAP XR 190 1345 1 1345 53 53 1087 1 26 27 87 Mar. 21, 2000 181 HFABH95 209407 Uni-ZAP XR 191 1347 1 1347 199 199 1088 1 21 22 116 Oct. 23, 1997 182 HFAMB72 209146 Uni-ZAP XR 192 1323 509 1323 559 559 1089 1 22 23 60 Jul. 17, 1997 183 HFAMH77 209300 Uni-ZAP XR 193 669 96 669 240 240 1090 1 33 34 61 Sep. 25, 1997 184 HFCCQ50 209463 Uni-ZAP XR 194 1271 1 1271 47 47 1091 1 20 21 352 Nov. 14, 1997 185 HFCDK17 97923 Uni-ZAP XR 195 1448 475 1392 567 567 1092 1 30 Mar. 07, 1997 209071 May 22, 1997 186 HFCEW05 209603 Uni-ZAP XR 196 933 1 933 34 34 1093 1 18 19 209 Jan. 29, 1998 187 HFFAD59 209242 Lambda ZAP II 197 470 1 470 44 44 1094 1 17 18 45 Sep. 12, 1997 188 HFFAL36 209368 Lambda ZAP II 198 1020 1 1020 68 68 1095 1 35 36 56 Oct. 16, 1997 189 HFGAD82 209225 Uni-ZAP XR 199 1881 772 1861 1019 1019 1096 1 18 19 38 Aug. 28, 1997 190 HFIIN69 PTA-846 pSport1 200 1450 1 1450 45 45 1097 1 39 40 43 Oct. 13, 1999 190 HFIIN69 PTA-846 pSport1 690 559 1 559 52 52 1587 1 39 40 43 Oct. 13, 1999 190 HFIIN69 PTA-846 pSport1 691 678 1 678 280 1588 1 2 Oct. 13, 1999 191 HFIIZ70 PTA-846 pSport1 201 1408 1 1408 24 24 1098 1 23 24 47 Oct. 13, 1999 191 HFIIZ70 PTA-846 pSport1 692 1441 43 1441 74 74 1589 1 23 24 47 Oct. 13, 1999 192 HFKET18 PTA-622 Uni-ZAP XR 202 2407 1 2407 137 137 1099 1 14 15 74 Sep. 02, 1999 193 HFLNB64 209463 Uni-ZAP XR 203 648 1 648 62 62 1100 1 39 40 45 Nov. 14, 1997 194 HFOXA73 209277 pSport1 204 540 1 540 25 25 1101 1 17 18 52 Sep. 18, 1997 194 HFOXA73 209277 pSport1 693 539 1 539 15 15 1590 1 17 Sep. 18, 1997 195 HFOXB13 209423 pSport1 205 1169 1 1169 36 36 1102 1 21 22 54 Oct. 30, 1997 196 HFPAC12 209511 Uni-ZAP XR 206 1088 1 1088 140 140 1103 1 21 22 88 Dec. 03, 1997 197 HFPAO71 209626 Uni-ZAP XR 207 2067 364 2067 414 414 1104 1 33 34 131 Feb. 12, 1998 198 HFPCX09 209551 Uni-ZAP XR 208 2213 1 2213 185 185 1105 1 26 27 549 Dec. 12, 1997 198 HFPCX09 209551 Uni-ZAP XR 694 2674 59 2674 249 249 1591 1 26 27 549 Dec. 12, 1997 198 HFPCX09 209551 Uni-ZAP XR 695 2207 1 2207 185 185 1592 1 26 27 66 Dec. 12, 1997 199 HFPCX36 209242 Uni-ZAP XR 209 796 1 796 103 103 1106 1 27 28 46 Sep. 12, 1997 200 HFPCX64 209878 Uni-ZAP XR 210 1076 1 1076 181 181 1107 1 28 29 87 May 18, 1998 200 HFPCX64 209878 Uni-ZAP XR 696 1069 1 1069 181 181 1593 1 28 29 181 May 18, 1998 200 HFPCX64 209878 Uni-ZAP XR 697 1154 84 1154 257 257 1594 1 28 29 87 May 18, 1998 200 HFPCX64 209878 Uni-ZAP XR 698 1197 85 1197 257 257 1595 1 28 29 87 May 18, 1998 201 HFRAN90 209242 Uni-ZAP XR 211 532 1 532 178 178 1108 1 33 34 54 Sep. 12, 1997 202 HFTBM50 209300 Uni-ZAP XR 212 762 1 740 158 158 1109 1 20 21 34 Sep. 25, 1997 203 HFTDL56 209782 Uni-ZAP XR 213 1839 32 1838 93 93 1110 1 20 21 519 Apr. 20, 1998 204 HFVAB79 209368 Uni-ZAP XR 214 1175 1 1175 133 133 1111 1 15 16 194 Oct. 16, 1997 204 HFVAB79 209368 Uni-ZAP XR 699 1186 1 1186 139 139 1596 1 15 16 194 Oct. 16, 1997 205 HFXAM76 209568 Lambda ZAP II 215 947 1 947 213 213 1112 1 24 25 79 Jan. 06, 1998 206 HFXDJ75 209603 Lambda ZAP II 216 1918 1 1914 44 44 1113 1 26 27 41 Jan. 29, 1998 207 HFXDN63 209346 Lambda ZAP II 217 1026 1 1026 33 33 1114 1 14 15 53 Oct. 09, 1997 208 HFXGT26 209965 Lambda ZAP II 218 1757 1 1757 13 13 1115 1 22 23 85 Jun. 11, 1998 209 HFXGV31 209242 Lambda ZAP II 219 752 1 752 100 100 1116 1 24 25 64 Sep. 12, 1997 210 HFXHD88 209511 Lambda ZAP II 220 1602 1 1602 130 130 1117 1 41 42 128 Dec. 03, 1997 211 HFXJU68 209423 Lambda ZAP II 221 712 1 712 141 141 1118 1 26 27 162 Oct. 30, 1997 211 HFXJU68 209423 Lambda ZAP II 700 1347 1 1347 148 148 1597 1 25 26 66 Oct. 30, 1997 212 HFXKJ03 209215 Lambda ZAP II 222 941 1 941 179 179 1119 1 33 34 41 Aug. 21, 1997 213 HFXKY27 209877 Lambda ZAP II 223 945 1 945 44 44 1120 1 19 20 58 May 18, 1998 214 HGBFO79 209011 Uni-ZAP XR 224 1538 259 1538 273 273 1121 1 23 24 49 Apr. 28, 1997 215 HGBHE57 209407 Uni-ZAP XR 225 663 1 663 14 14 1122 1 19 20 68 Oct. 23, 1997 216 HGBIB74 203648 Uni-ZAP XR 226 1816 1 1804 14 14 1123 1 23 24 377 Feb. 09, 1999 216 HGBIB74 203648 Uni-ZAP XR 701 1821 1 1821 28 28 1598 1 20 21 170 Feb. 09, 1999 216 HGBIB74 203648 Uni-ZAP XR 702 1094 1 1094 2 1599 1 1 2 151 Feb. 09, 1999 217 HGLAL82 209242 Uni-ZAP XR 227 406 1 406 144 144 1124 1 19 20 26 Sep. 12, 1997 218 HHAAF20 203648 Uni-ZAP XR 228 1495 1 1495 141 141 1125 1 18 19 55 Feb. 09, 1999 219 HHEAA08 209853 pCMVSport 3.0 229 2150 1 2150 88 88 1126 1 38 39 79 May 07, 1998 219 HHEAA08 209853 pCMVSport 3.0 703 615 1 615 311 1600 1 13 14 20 May 07, 1998 220 HHEBB10 209568 pCMVSport 3.0 230 1827 141 1810 334 334 1127 1 23 24 99 Jan. 06, 1998 221 HHEMA59 203364 pCMVSport 3.0 231 3102 1 3099 239 239 1128 1 20 21 76 Oct. 19, 1998 222 HHEMA75 209179 pCMVSport 3.0 232 865 229 865 569 569 1129 1 35 36 84 Jul. 24, 1997 223 HHEMM74 PTA-849 pCMVSport 3.0 233 2612 1 2612 94 94 1130 1 27 28 74 Oct. 13, 1999 223 HHEMM74 PTA-849 pCMVSport 3.0 704 1125 1 1125 121 121 1601 1 27 28 74 Oct. 13, 1999 223 HHEMM74 PTA-849 pCMVSport 3.0 705 2297 1425 2297 706 1602 1 6 7 33 Oct. 13, 1999 223 HHEMM74 PTA-849 pCMVSport 3.0 706 482 33 482 7 1603 1 13 14 53 Oct. 13, 1999 224 HHENK42 209195 pCMVSport 3.0 234 656 1 656 63 63 1131 1 7 8 42 Aug. 01, 1997 225 HHENP27 203105 pCMVSport 3.0 235 1237 1 1237 12 12 1132 1 22 23 282 Aug. 13, 1998 226 HHENQ22 209511 pCMVSport 3.0 236 1899 1 1899 115 115 1133 1 36 37 58 Dec. 03, 1997 227 HHEPD24 209195 pCMVSport 3.0 237 238 1 238 156 156 1134 1 23 24 27 Aug. 01, 1997 228 HHEPM33 PTA-322 pCMVSport 3.0 238 1459 1 1459 269 269 1135 1 20 21 82 Jul. 09, 1999 229 HHEPT60 209138 pCMVSport 3.0 239 532 21 532 245 245 1136 1 18 19 36 Jul. 03, 1997 230 HHEPU04 203648 pCMVSport 3.0 240 1084 116 1084 259 259 1137 1 31 32 163 Feb. 09, 1999 230 HHEPU04 203648 pCMVSport 3.0 707 1081 124 1081 267 267 1604 1 31 32 163 Feb. 09, 1999 230 HHEPU04 203648 pCMVSport 3.0 708 720 1 720 45 45 1605 1 31 32 92 Feb. 09, 1999 231 HHFEC49 PTA-844 Uni-ZAP XR 241 2263 1 2263 30 30 1138 1 24 25 184 Oct. 13, 1999 232 HHFGR93 209746 Uni-ZAP XR 242 1835 1 1835 132 132 1139 1 29 30 390 Apr. 07, 1998 232 HHFGR93 209746 Uni-ZAP XR 709 1932 1 1836 130 130 1606 1 29 30 236 Apr. 07, 1998 233 HHFHJ59 97975 Uni-ZAP XR 243 661 1 661 192 192 1140 1 29 30 112 Apr. 04, 1997 209081 May 29, 1997 234 HHFHR32 97975 Uni-ZAP XR 244 1378 1 1378 58 58 1141 1 25 26 235 Apr. 04, 1997 209081 May 29, 1997 235 HHFOJ29 PTA-2075 Uni-ZAP XR 245 1366 1 1366 117 117 1142 1 31 32 82 Jun. 09, 2000 235 HHFOJ29 PTA-2075 Uni-ZAP XR 710 1595 513 1595 132 132 1607 1 19 20 95 Jun. 09, 2000 235 HHFOJ29 PTA-2075 Uni-ZAP XR 711 970 272 970 62 1608 1 1 2 152 Jun. 09, 2000 236 HHGCM76 97958 Lambda ZAP II 246 711 8 711 270 270 1143 1 22 23 89 Mar. 13, 1997 209072 May 22, 1997 236 HHGCM76 97958 Lambda ZAP II 712 711 8 711 270 270 1609 1 11 Mar. 13, 1997 209072 May 22, 1997 237 HHGDF16 209463 Lambda ZAP II 247 890 215 890 253 253 1144 1 26 27 52 Nov. 14, 1997 238 HHGDW43 209346 Lambda ZAP II 248 1050 1 1050 107 107 1145 1 40 41 44 Oct. 09, 1997 239 HHPEC09 209877 Uni-ZAP XR 249 488 1 488 71 71 1146 1 19 20 55 May 18, 1998 240 HHPGO40 209878 Uni-ZAP XR 250 1002 1 1002 116 116 1147 1 26 27 295 May 18, 1998 240 HHPGO40 209878 Uni-ZAP XR 713 973 1 973 68 68 1610 1 37 38 302 May 18, 1998 240 HHPGO40 209878 Uni-ZAP XR 714 984 1 984 74 74 1611 1 37 38 224 May 18, 1998 241 HHPTJ65 209179 Uni-ZAP XR 251 515 1 515 247 247 1148 1 32 33 48 Jul. 24, 1997 242 HHSDX28 209346 Uni-ZAP XR 252 1113 1 1113 90 90 1149 1 21 22 56 Oct. 09, 1997 243 HHSGW69 PTA-855 Uni-ZAP XR 253 1254 1 1254 238 238 1150 1 26 27 55 Oct. 18, 1999 243 HHSGW69 PTA-855 Uni-ZAP XR 715 826 1 826 231 231 1612 1 26 27 55 Oct. 18, 1999 243 HHSGW69 PTA-855 Uni-ZAP XR 716 4400 1605 1674 457 1613 1 1 2 314 Oct. 18, 1999 244 HHTLF25 209125 ZAP Express 254 697 1 661 142 142 1151 1 26 27 111 Jun. 19, 1997 245 HJABX32 209146 pBLUESCRIPT ™ 255 1061 454 1061 557 557 1152 1 18 19 51 Jul. 17, 1997 SK− 246 HJACA79 209368 pBLUESCRIPT ™ 256 887 1 887 84 84 1153 1 28 29 68 Oct. 16, 1997 SK− 247 HJACG02 209215 pBLUESCRIPT ™ 257 575 1 575 66 66 1154 1 22 23 108 Aug. 21, 1997 SK− 247 HJACG02 209215 pBLUESCRIPT ™ 717 553 1 553 47 47 1614 1 23 24 108 Aug. 21, 1997 SK− 248 HJACG30 PTA-843 pBLUESCRIPT ™ 258 1532 1 1532 291 291 1155 1 27 28 44 Oct. 13, 1999 SK− 248 HJACG30 PTA-843 pBLUESCRIPT ™ 718 1614 1020 1614 50 1615 1 1 2 130 Oct. 13, 1999 SK− 248 HJACG30 PTA-843 pBLUESCRIPT ™ 719 1087 491 1087 350 1616 1 1 2 122 Oct. 13, 1999 SK− 249 HJBAV55 203364 pBLUESCRIPT ™ 259 2441 39 2429 238 238 1156 1 26 27 58 Oct. 19, 1998 SK− 250 HJBCU04 PTA-322 pBLUESCRIPT ™ 260 1192 1 1192 96 96 1157 1 49 50 176 Jul. 09, 1999 SK− 251 HJMBI18 209580 pCMVSport 3.0 261 1021 303 1021 574 574 1158 1 19 20 80 Jan. 14, 1998 252 HJMBN89 209407 pCMVSport 3.0 262 1064 306 1064 348 348 1159 1 13 14 56 Oct. 23, 1997 253 HJMBT65 209580 pCMVSport 3.0 263 621 79 621 341 341 1160 1 33 34 42 Jan. 14, 1998 254 HJMBW30 209146 pCMVSport 3.0 264 884 1 874 110 110 1161 1 18 19 42 Jul. 17, 1997 255 HJPAD75 209641 Uni-ZAP XR 265 1231 1 1231 60 60 1162 1 29 30 91 Feb. 25, 1998 256 HKAAE44 209368 pCMVSport 2.0 266 1494 1 1494 113 113 1163 1 39 40 136 Oct. 16, 1997 257 HKAAH36 209563 pCMVSport 2.0 267 1216 1 1216 128 128 1164 1 29 30 293 Dec. 18, 1997 257 HKAAH36 209563 pCMVSport 2.0 720 1016 1 1016 295 295 1617 1 29 30 143 Dec. 18, 1997 257 HKAAH36 209563 pCMVSport 2.0 721 1490 1 1490 182 182 1618 1 29 30 293 Dec. 18, 1997 257 HKAAH36 209563 pCMVSport 2.0 722 1441 8 1392 184 184 1619 1 29 30 85 Dec. 18, 1997 257 HKAAH36 209563 pCMVSport 2.0 723 1516 1 1516 254 254 1620 1 29 30 293 Dec. 18, 1997 257 HKAAH36 209563 pCMVSport 2.0 724 1381 196 1381 129 129 1621 1 29 30 293 Dec. 18, 1997 257 HKAAH36 209563 pCMVSport 2.0 725 1439 1 1439 189 189 1622 1 29 30 61 Dec. 18, 1997 258 HKAAK02 209551 pCMVSport 2.0 268 859 1 859 97 97 1165 1 34 35 196 Dec. 12, 1997 259 HKABI84 209603 pCMVSport 2.0 269 1238 45 1238 274 274 1166 1 16 17 47 Jan. 29, 1998 260 HKABZ65 209683 pCMVSport 2.0 270 1189 1 1189 77 77 1167 1 17 18 243 Mar. 20, 1998 260 HKABZ65 209683 pCMVSport 2.0 726 1191 1 1191 69 69 1623 1 17 18 243 Mar. 20, 1998 261 HKACB56 209346 pCMVSport 2.0 271 496 1 496 27 27 1168 1 23 24 80 Oct. 09, 1997 262 HKACD58 209346 pCMVSport 2.0 272 3153 1 3153 38 38 1169 1 25 26 301 Oct. 09, 1997 262 HKACD58 209346 pCMVSport 2.0 727 1626 1 1626 35 35 1624 1 25 26 154 Oct. 09, 1997 263 HKACM93 PTA-849 pCMVSport 2.0 273 2352 1 2352 218 218 1170 1 30 31 692 Oct. 13, 1999 263 HKACM93 PTA-849 pCMVSport 2.0 728 549 1 549 189 189 1625 1 30 31 120 Oct. 13, 1999 263 HKACM93 PTA-849 pCMVSport 2.0 729 1120 1 1120 314 314 1626 1 30 31 269 Oct. 13, 1999 263 HKACM93 PTA-849 pCMVSport 2.0 730 1893 739 1893 202 1627 1 13 14 17 Oct. 13, 1999 263 HKACM93 PTA-849 pCMVSport 2.0 731 1187 1 1187 638 1628 1 4 5 45 Oct. 13, 1999 264 HKADQ91 209568 pCMVSport 2.0 274 1523 30 1517 229 229 1171 1 25 26 275 Jan. 06, 1998 265 HKAEG43 209965 pCMVSport 2.0 275 1297 1 1297 32 32 1172 1 29 30 70 Jun. 11, 1998 265 HKAEG43 209965 pCMVSport 2.0 732 1286 1 1286 21 21 1629 1 29 30 70 Jun. 11, 1998 266 HKAEL80 209423 pCMVSport 2.0 276 1105 1 1105 398 398 1173 1 17 18 79 Oct. 30, 1997 267 HKAEV06 209627 pCMVSport 2.0 277 2496 1 2496 501 501 1174 1 30 31 438 Feb. 12, 1998 267 HKAEV06 209627 pCMVSport 2.0 733 2351 1 2351 197 197 1630 1 29 30 57 Feb. 12, 1998 268 HKAFK41 209300 pCMVSport 2.0 278 549 1 549 243 243 1175 1 30 31 43 Sep. 25, 1997 269 HKDBF34 209511 pCMVSport 1 279 1432 60 1418 69 69 1176 1 14 15 222 Dec. 03, 1997 269 HKDBF34 209511 pCMVSport 1 734 1356 1 1356 18 18 1631 1 19 20 104 Dec. 03, 1997 270 HKGAT94 209126 pSport1 280 1048 1 1048 449 449 1177 1 31 32 99 Jun. 19, 1997 270 HKGAT94 209126 pSport1 735 1063 1 1063 470 1632 1 20 21 94 Jun. 19, 1997 271 HKGCO27 209853 pSport1 281 1021 1 1021 313 313 1178 1 26 27 93 May 07, 1998 271 HKGCO27 209853 pSport1 736 1311 1 1311 57 57 1633 1 26 27 47 May 07, 1998 272 HKISB57 209603 pBLUESCRIPT ™ 282 1492 1 1439 130 130 1179 1 19 20 95 Jan. 29, 1998 273 HKIYH57 209324 pBLUESCRIPT ™ 283 609 156 609 336 336 1180 1 23 24 54 Oct. 02, 1997 274 HKIYP40 209463 pBLUESCRIPT ™ 284 1215 1 1215 43 43 1181 1 32 33 76 Nov. 14, 1997 275 HKMLK53 209511 pBLUESCRIPT ™ 285 1543 1 1543 20 20 1182 1 25 26 69 Dec. 03, 1997 276 HKMLP68 PTA-845 pBLUESCRIPT ™ 286 2784 1 2784 130 130 1183 1 24 25 80 Oct. 13, 1999 276 HKMLP68 PTA-845 pBLUESCRIPT ™ 737 718 1 718 153 153 1634 1 24 25 80 Oct. 13, 1999 276 HKMLP68 PTA-845 pBLUESCRIPT ™ 738 614 1 614 471 1635 1 1 2 47 Oct. 13, 1999 277 HL2AC08 209580 Uni-ZAP XR 287 1478 1 1478 64 64 1184 1 23 24 280 Jan. 14, 1998 278 HL2AG57 209746 Uni-ZAP XR 288 1780 349 1780 560 560 1185 1 31 32 80 Apr. 07, 1998 279 HLCND09 PTA-2076 Uni-ZAP XR 289 1984 1 1984 146 146 1186 1 38 39 110 Jun. 09, 2000 279 HLCND09 PTA-2076 Uni-ZAP XR 739 465 1 465 38 38 1636 1 38 39 142 Jun. 09, 2000 280 HLDBX13 203331 pCMVSport 3.0 290 1815 1 1815 303 303 1187 1 39 40 55 Oct. 08, 1998 281 HLDON23 209628 pCMVSport 3.0 291 1262 208 1256 368 368 1188 1 20 21 113 Feb. 12, 1998 282 HLDOW79 PTA-1544 pCMVSport 3.0 292 989 1 989 43 43 1189 1 21 22 275 Mar. 21, 2000 283 HLDQC46 PTA-1544 pCMVSport 3.0 293 632 1 632 163 163 1190 1 34 35 87 Mar. 21, 2000 284 HLDQR62 203027 pCMVSport 3.0 294 2572 427 2572 520 520 1191 1 18 19 161 Jun. 26, 1998 285 HLDQU79 203071 pCMVSport 3.0 295 1488 1 1488 99 99 1192 1 23 24 348 Jul. 27, 1998 286 HLDRM43 209628 pCMVSport 3.0 296 609 1 609 24 24 1193 1 20 21 151 Feb. 12, 1998 286 HLDRM43 209628 pCMVSport 3.0 740 759 1 759 164 164 1637 1 20 21 151 Feb. 12, 1998 287 HLDRP33 209641 pCMVSport 3.0 297 612 1 612 215 215 1194 1 26 27 41 Feb. 25, 1998 288 HLHFP03 209126 Uni-ZAP XR 298 613 1 613 224 224 1195 1 19 20 116 Jun. 19, 1997 289 HLHFR58 PTA-841 Uni-ZAP XR 299 1015 1 1015 206 1196 1 17 18 21 Oct. 13, 1999 289 HLHFR58 PTA-841 Uni-ZAP XR 741 733 1 733 205 1638 1 16 17 21 Oct. 13, 1999 289 HLHFR58 PTA-841 Uni-ZAP XR 742 741 1 741 288 1639 1 1 2 67 Oct. 13, 1999 289 HLHFR58 PTA-841 Uni-ZAP XR 743 951 12 675 254 1640 1 1 2 91 Oct. 13, 1999 290 HLIBD68 203071 pCMVSport 1 300 1022 1 1022 186 186 1197 1 35 36 50 Jul. 27, 1998 291 HLICQ90 203517 pCMVSport 1 301 1766 1 1766 249 249 1198 1 29 30 206 Dec. 10, 1998 292 HLJBJ61 PTA-848 pCMVSport 1 302 1191 1 1191 158 158 1199 1 29 30 38 Oct. 13, 1999 292 HLJBJ61 PTA-848 pCMVSport 1 744 628 1 628 227 227 1641 1 29 30 38 Oct. 13, 1999 293 HLMBO76 209603 Lambda ZAP II 303 815 1 795 43 43 1200 1 43 44 107 Jan. 29, 1998 294 HLMCA59 209236 Uni-ZAP XR 304 787 1 787 101 101 1201 1 31 32 63 Sep. 04, 1997 295 HLQBE09 209243 Lambda ZAP II 305 633 1 633 17 17 1202 1 19 20 181 Sep. 12, 1997 296 HLQDH79 209551 Lambda ZAP II 306 913 1 913 205 205 1203 1 19 20 58 Dec. 12, 1997 297 HLQDR48 209603 Lambda ZAP II 307 989 1 989 10 10 1204 1 21 22 190 Jan. 29, 1998 297 HLQDR48 209603 Lambda ZAP II 745 990 1 990 3 3 1642 1 21 22 190 Jan. 29, 1998 298 HLQEM64 PTA-623 Lambda ZAP II 308 774 1 774 247 247 1205 1 29 30 144 Sep. 02, 1999 298 HLQEM64 PTA-623 Lambda ZAP II 746 1038 1 702 42 42 1643 1 29 30 132 Sep. 02, 1999 299 HLTAU74 PTA-163 Uni-ZAP XR 309 1524 1 1524 76 76 1206 1 21 22 62 Jun. 01, 1999 300 HLTCO33 203071 Uni-ZAP XR 310 1184 1 1184 80 80 1207 1 18 19 64 Jul. 27, 1998 301 HLTDV50 209243 Uni-ZAP XR 311 770 1 770 74 74 1208 1 17 18 28 Sep. 12, 1997 302 HLTEJ06 209346 Uni-ZAP XR 312 617 69 617 197 197 1209 1 22 23 55 Oct. 09, 1997 303 HLTFA64 209628 Uni-ZAP XR 313 1130 1 1130 268 268 1210 1 42 43 43 Feb. 12, 1998 304 HLTHG37 209965 Uni-ZAP XR 314 3740 1908 3740 50 50 1211 1 1 2 319 Jun. 11, 1998 304 HLTHG37 209965 Uni-ZAP XR 747 1932 98 1932 313 313 1644 1 35 36 42 Jun. 11, 1998 305 HLWAA17 209626 pCMVSport 3.0 315 997 246 997 436 436 1212 1 15 16 187 Feb. 12, 1998 306 HLWAD77 209651 pCMVSport 3.0 316 1167 304 1167 326 326 1213 1 24 25 140 Mar. 04, 1998 307 HLWAE11 203071 pCMVSport 3.0 317 1618 1 1618 28 28 1214 1 46 47 278 Jul. 27, 1998 308 HLWAO22 209511 pCMVSport 3.0 318 1338 1 1311 212 212 1215 1 21 22 354 Dec. 03, 1997 309 HLWAY54 209651 pCMVSport 3.0 319 1892 1 1892 38 38 1216 1 25 26 338 Mar. 04, 1998 310 HLWBI63 209407 pCMVSport 3.0 320 1038 1 1038 149 149 1217 1 30 31 63 Oct. 23, 1997 311 HLWBY76 203517 pCMVSport 3.0 321 2081 1 2081 432 432 1218 1 27 28 232 Dec. 10, 1998 312 HLWCF05 209126 pCMVSport 3.0 322 646 1 646 155 155 1219 1 36 37 58 Jun. 19, 1997 313 HLYAC95 203071 pSport1 323 312 1 312 92 92 1220 1 16 17 46 Jul. 27, 1998 314 HLYAF80 209126 pSport1 324 826 1 826 222 222 1221 1 24 25 47 Jun. 19, 1997 315 HLYAN59 209346 pSport1 325 770 1 770 383 383 1222 1 40 41 77 Oct. 09, 1997 315 HLYAN59 209346 pSport1 748 729 1 729 254 254 1645 1 39 40 54 Oct. 09, 1997 316 HLYAZ61 209022 pSport1 326 1237 1 1237 190 190 1223 1 18 19 222 May 08, 1997 316 HLYAZ61 209022 pSport1 749 997 74 997 205 205 1646 1 18 19 215 May 08, 1997 317 HLYBD32 209407 pSport1 327 1045 35 1045 98 98 1224 1 23 24 70 Oct. 23, 1997 318 HMADS41 209563 Uni-ZAP XR 328 1267 1 1267 267 267 1225 1 21 22 88 Dec. 18, 1997 319 HMADU73 209139 Uni-ZAP XR 329 3194 1 3194 491 491 1226 1 16 17 713 Jul. 03, 1997 319 HMADU73 209139 Uni-ZAP XR 750 437 1 437 115 115 1647 1 15 16 77 Jul. 03, 1997 320 HMAMI15 PTA-2075 Uni-ZAP XR 330 1258 1 1258 4 4 1227 1 26 27 340 Jun. 09, 2000 320 HMAMI15 PTA-2075 Uni-ZAP XR 751 1084 1 1084 3 3 1648 1 26 27 306 Jun. 09, 2000 321 HMDAE65 209243 Uni-ZAP XR 331 698 1 698 179 179 1228 1 17 18 77 Sep. 12, 1997 322 HMDAN54 97974 Uni-ZAP XR 332 1856 725 1853 928 928 1229 1 33 34 50 Apr. 04, 1997 209080 May 29, 1997 323 HMDAQ29 209563 Uni-ZAP XR 333 974 1 974 180 180 1230 1 43 44 82 Dec. 18, 1997 324 HMEAI48 203069 Lambda ZAP II 334 413 1 413 36 36 1231 1 29 30 88 Jul. 27, 1998 324 HMEAI48 203069 Lambda ZAP II 752 1168 1 1168 95 95 1649 1 29 30 40 Jul. 27, 1998 325 HMECK83 209853 Lambda ZAP II 335 1010 1 1010 50 50 1232 1 28 29 54 May 07, 1998 326 HMEED18 209368 Lambda ZAP II 336 1369 28 1369 34 34 1233 1 34 35 221 Oct. 16, 1997 327 HMEET96 209407 Lambda ZAP II 337 1337 73 1200 121 121 1234 1 30 31 266 Oct. 23, 1997 328 HMIAL37 209563 Uni-ZAP XR 338 1420 1 1420 49 49 1235 1 13 14 97 Dec. 18, 1997 329 HMIAP86 209878 Uni-ZAP XR 339 1674 13 1674 182 182 1236 1 19 20 334 May 18, 1998 330 HMKCG09 209346 pSport1 340 921 60 921 221 221 1237 1 28 29 49 Oct. 09, 1997 331 HMMAH60 209368 pSport1 341 822 1 822 142 142 1238 1 15 16 50 Oct. 16, 1997 332 HMQDF12 209407 Uni-ZAP XR 342 706 1 627 63 63 1239 1 27 28 142 Oct. 23, 1997 333 HMQDT36 209022 Uni-ZAP XR 343 1871 1 1871 157 157 1240 1 32 33 406 May 08, 1997 333 HMQDT36 209022 Uni-ZAP XR 753 1914 37 1897 192 192 1650 1 32 33 406 May 08, 1997 334 HMSBX80 209563 Uni-ZAP XR 344 1726 1 1726 169 169 1241 1 19 20 57 Dec. 18, 1997 335 HMSFS21 209324 Uni-ZAP XR 345 1283 1 1283 28 28 1242 1 17 18 37 Oct. 02, 1997 336 HMSGB14 209423 Uni-ZAP XR 346 1552 1 1552 138 138 1243 1 18 19 77 Oct. 30, 1997 337 HMSGU01 PTA-1544 Uni-ZAP XR 347 1617 1 1617 137 137 1244 1 23 24 120 Mar. 21, 2000 337 HMSGU01 PTA-1544 Uni-ZAP XR 754 1257 1 1257 137 137 1651 1 23 24 235 Mar. 21, 2000 337 HMSGU01 PTA-1544 Uni-ZAP XR 755 1654 1 1654 135 135 1652 1 23 24 120 Mar. 21, 2000 338 HMSHM14 209126 Uni-ZAP XR 348 756 1 756 103 103 1245 1 29 30 45 Jun. 19, 1997 339 HMSHS36 PTA-2070 Uni-ZAP XR 349 1402 1 1402 134 134 1246 1 23 24 103 Jun. 09, 2000 339 HMSHS36 PTA-2070 Uni-ZAP XR 756 616 30 616 162 162 1653 1 23 24 103 Jun. 09, 2000 340 HMSJM65 209641 Uni-ZAP XR 350 2270 1 2231 111 111 1247 1 27 28 77 Feb. 25, 1998 341 HMSJU68 209076 Uni-ZAP XR 351 1123 4 1123 272 272 1248 1 31 32 49 May 22, 1997 342 HMSKC04 203105 Uni-ZAP XR 352 1417 1 1417 133 133 1249 1 22 23 73 Aug. 13, 1998 343 HMTAD67 209551 pCMVSport 3.0 353 1173 1 1173 306 306 1250 1 19 20 84 Dec. 12, 1997 344 HMUAP70 209878 pCMVSport 3.0 354 1965 531 1914 183 183 1251 1 16 17 221 May 18, 1998 344 HMUAP70 209878 pCMVSport 3.0 757 1842 407 1783 413 413 1654 1 25 26 103 May 18, 1998 344 HMUAP70 209878 pCMVSport 3.0 758 1963 530 1914 251 251 1655 1 28 29 198 May 18, 1998 344 HMUAP70 209878 pCMVSport 3.0 759 1487 1 1487 62 62 1656 1 16 17 106 May 18, 1998 344 HMUAP70 209878 pCMVSport 3.0 760 1653 1 1653 60 60 1657 1 15 16 68 May 18, 1998 344 HMUAP70 209878 pCMVSport 3.0 761 1830 407 1830 60 60 1658 1 23 May 18, 1998 345 HMVBN46 209603 pSport1 355 1382 1 1382 10 10 1252 1 19 20 48 Jan. 29, 1998 346 HMWEB02 209628 Uni-ZAP XR 356 1755 1 1755 106 106 1253 1 23 24 91 Feb. 12, 1998 347 HMWFO02 209324 Uni-ZAP XR 357 547 1 547 7 7 1254 1 37 38 68 Oct. 02, 1997 347 HMWFO02 209324 Uni-ZAP XR 762 708 1 708 20 20 1659 1 38 39 60 Oct. 02, 1997 348 HMWFY10 209147 Uni-ZAP XR 358 556 1 556 367 367 1255 1 15 16 30 Jul. 17, 1997 348 HMWFY10 209147 Uni-ZAP XR 763 556 1 556 129 1660 1 9 10 18 Jul. 17, 1997 349 HMWGY65 203105 Uni-ZAP XR 359 1974 1 1974 42 42 1256 1 21 22 490 Aug. 13, 1998 349 HMWGY65 203105 Uni-ZAP XR 764 2027 1 1976 42 42 1661 1 21 22 188 Aug. 13, 1998 350 HNEAC05 209236 Uni-ZAP XR 360 890 1 890 101 101 1257 1 24 25 105 Sep. 04, 1997 351 HNEEB45 PTA-845 Uni-ZAP XR 361 1043 1 1043 139 139 1258 1 25 26 57 Oct. 13, 1999 351 HNEEB45 PTA-845 Uni-ZAP XR 765 699 160 699 226 226 1662 1 25 26 57 Oct. 13, 1999 352 HNFFC43 203027 Uni-ZAP XR 362 2103 209 2058 488 488 1259 1 12 13 68 Jun. 26, 1998 353 HNFGF20 203071 Uni-ZAP XR 363 1370 38 1370 206 206 1260 1 45 46 143 Jul. 27, 1998 354 HNFJF07 209463 Uni-ZAP XR 364 616 1 616 86 86 1261 1 21 22 66 Nov. 14, 1997 355 HNFJH45 97976 Uni-ZAP XR 365 575 1 575 275 275 1262 1 30 31 67 Apr. 04, 1997 356 HNGAK47 209368 Uni-ZAP XR 366 1144 1 1144 89 89 1263 1 23 24 40 Oct. 16, 1997 357 HNGAP93 209243 Uni-ZAP XR 367 703 1 703 50 50 1264 1 19 20 33 Sep. 12, 1997 358 HNGBC07 PTA-844 Uni-ZAP XR 368 1649 1 1647 81 81 1265 1 18 19 249 Oct. 13, 1999 358 HNGBC07 PTA-844 Uni-ZAP XR 766 1649 1 1647 122 122 1663 1 24 25 44 Oct. 13, 1999 358 HNGBC07 PTA-844 Uni-ZAP XR 767 1570 1 1570 55 55 1664 1 24 25 44 Oct. 13, 1999 359 HNGBT31 97976 Uni-ZAP XR 369 639 1 639 224 224 1266 1 28 29 104 Apr. 04, 1997 360 HNGDJ72 209299 Uni-ZAP XR 370 524 1 524 185 185 1267 1 19 20 113 Sep. 25, 1997 361 HNGDU40 209563 Uni-ZAP XR 371 1035 1 1035 333 333 1268 1 17 18 51 Dec. 18, 1997 362 HNGEG08 209179 Uni-ZAP XR 372 660 1 660 94 94 1269 1 35 36 66 Jul. 24, 1997 363 HNGEO29 209299 Uni-ZAP XR 373 491 1 491 98 98 1270 1 32 33 44 Sep. 25, 1997 364 HNGEP09 209197 Uni-ZAP XR 374 1042 1 1042 72 72 1271 1 15 16 82 Aug. 08, 1997 365 HNGHR74 209346 Uni-ZAP XR 375 1095 1 1095 53 53 1272 1 18 19 41 Oct. 09, 1997 366 HNGIH43 97976 Uni-ZAP XR 376 427 1 427 178 178 1273 1 31 32 40 Apr. 04, 1997 367 HNGIJ31 209236 Uni-ZAP XR 377 796 1 796 135 135 1274 1 16 17 36 Sep. 04, 1997 368 HNGIQ46 209243 Uni-ZAP XR 378 527 1 527 221 221 1275 1 21 22 70 Sep. 12, 1997 369 HNGJE50 209368 Uni-ZAP XR 379 1037 1 1037 77 77 1276 1 36 37 46 Oct. 16, 1997 370 HNGJO57 209463 Uni-ZAP XR 380 828 1 828 87 87 1277 1 18 19 52 Nov. 14, 1997 371 HNGJP69 209603 Uni-ZAP XR 381 985 1 985 321 321 1278 1 14 15 74 Jan. 29, 1998 372 HNGJT54 209215 Uni-ZAP XR 382 1110 1 1110 172 172 1279 1 19 20 34 Aug. 21, 1997 373 HNGOI12 PTA-847 Uni-ZAP XR 383 2128 1 2128 27 27 1280 1 34 35 57 Oct. 13, 1999 373 HNGOI12 PTA-847 Uni-ZAP XR 768 774 1 774 27 27 1665 1 34 35 57 Oct. 13, 1999 373 HNGOI12 PTA-847 Uni-ZAP XR 769 1396 1 1396 596 1666 1 25 26 93 Oct. 13, 1999 374 HNGOM56 203648 Uni-ZAP XR 384 956 1 956 391 391 1281 1 22 23 55 Feb. 09, 1999 375 HNHAH01 209180 Uni-ZAP XR 385 905 1 905 328 328 1282 1 41 42 54 Jul. 24, 1997 376 HNHCX60 209243 Uni-ZAP XR 386 762 1 762 158 158 1283 1 20 21 21 Sep. 12, 1997 377 HNHCY64 209243 Uni-ZAP XR 387 725 1 725 258 258 1284 1 32 33 44 Sep. 12, 1997 378 HNHCY94 209243 Uni-ZAP XR 388 606 1 606 78 78 1285 1 25 26 48 Sep. 12, 1997 379 HNHDW38 209299 Uni-ZAP XR 389 793 1 793 231 231 1286 1 22 23 46 Sep. 25, 1997 380 HNHDW42 97976 Uni-ZAP XR 390 426 1 426 168 168 1287 1 26 27 71 Apr. 04, 1997 381 HNHED17 209346 Uni-ZAP XR 391 843 1 843 274 274 1288 1 19 20 51 Oct. 09, 1997 381 HNHED17 209346 Uni-ZAP XR 770 692 1 692 282 282 1667 1 19 20 48 Oct. 09, 1997 382 HNHEI42 PTA-844 Uni-ZAP XR 392 2642 1 2642 52 52 1289 1 22 23 36 Oct. 13, 1999 382 HNHEI42 PTA-844 Uni-ZAP XR 771 1654 1 1654 28 28 1668 1 22 23 36 Oct. 13, 1999 382 HNHEI42 PTA-844 Uni-ZAP XR 772 447 1 447 166 1669 1 6 7 28 Oct. 13, 1999 382 HNHEI42 PTA-844 Uni-ZAP XR 773 641 1 641 331 1670 1 3 4 34 Oct. 13, 1999 383 HNHFO29 209138 Uni-ZAP XR 393 699 1 699 160 160 1290 1 21 22 180 Jul. 03, 1997 384 HNHFU32 209407 Uni-ZAP XR 394 607 1 607 175 175 1291 1 30 31 52 Oct. 23, 1997 385 HNHOD46 PTA-1543 Uni-ZAP XR 395 1355 1 1355 12 12 1292 1 20 21 80 Mar. 21, 2000 386 HNHOG73 203570 Uni-ZAP XR 396 802 1 802 342 342 1293 1 19 20 51 Jan. 11, 1999 387 HNTBL27 209324 pCMVSport 3.0 397 791 71 791 100 100 1294 1 23 24 115 Oct. 02, 1997 388 HNTCE26 PTA-1544 pCMVSport 3.0 398 2163 830 2163 111 111 1295 1 30 31 402 Mar. 21, 2000 388 HNTCE26 PTA-1544 pCMVSport 3.0 774 1763 1 1763 57 57 1671 1 28 29 121 Mar. 21, 2000 389 HNTNI01 209782 pSport1 399 2087 1 2087 307 307 1296 1 33 34 76 Apr. 20, 1998 389 HNTNI01 209782 pSport1 775 1274 1 1114 306 306 1672 1 33 34 49 Apr. 20, 1998 390 HOAAC90 209236 Uni-ZAP XR 400 642 1 642 33 33 1297 1 15 16 104 Sep. 04, 1997 390 HOAAC90 209236 Uni-ZAP XR 776 652 1 652 38 38 1673 1 15 16 104 Sep. 04, 1997 391 HOACB38 209243 Uni-ZAP XR 401 606 1 606 63 63 1298 1 21 22 40 Sep. 12, 1997 392 HOCNF19 203570 pSport1 402 1118 1 1118 166 166 1299 1 20 21 87 Jan. 11, 1999 393 HODDN65 209244 Uni-ZAP XR 403 755 1 755 251 251 1300 1 14 15 20 Sep. 12, 1997 394 HODDN92 209012 Uni-ZAP XR 404 1939 294 1939 434 1301 1 26 27 35 Apr. 28, 1997 209089 Jun. 05, 1997 395 HODDO08 203364 Uni-ZAP XR 405 1776 138 1284 725 725 1302 1 33 34 106 Oct. 19, 1998 396 HODDW40 209463 Uni-ZAP XR 406 682 1 682 139 139 1303 1 19 20 40 Nov. 14, 1997 397 HODFN71 203570 Uni-ZAP XR 407 1126 1 1126 1 1304 1 1 2 159 Jan. 11, 1999 397 HODFN71 203570 Uni-ZAP XR 777 1124 1 1124 27 27 1674 1 18 19 148 Jan. 11, 1999 398 HODGE68 203570 Uni-ZAP XR 408 851 1 851 87 87 1305 1 26 27 59 Jan. 11, 1999 399 HOEBK34 209224 Uni-ZAP XR 409 747 75 747 149 149 1306 1 20 21 165 Aug. 28, 1997 399 HOEBK34 209224 Uni-ZAP XR 778 660 1 660 68 68 1675 1 26 27 88 Aug. 28, 1997 400 HOEBZ89 203517 Uni-ZAP XR 410 2520 1 2520 19 19 1307 1 21 22 333 Dec. 10, 1998 401 HOEDB32 209628 Uni-ZAP XR 411 1462 73 1462 104 104 1308 1 21 22 226 Feb. 12, 1998 402 HOEDE28 PTA-844 Uni-ZAP XR 412 1635 1 1635 248 248 1309 1 21 22 117 Oct. 13, 1999 402 HOEDE28 PTA-844 Uni-ZAP XR 779 1424 806 1424 387 1676 1 11 12 20 Oct. 13, 1999 403 HOEDH84 209965 Uni-ZAP XR 413 2079 1 2079 256 256 1310 1 20 21 404 Jun. 11, 1998 404 HOFMQ33 PTA-848 pCMVSport 2.0 414 2410 1 2410 49 49 1311 1 24 25 484 Oct. 13, 1999 404 HOFMQ33 PTA-848 pCMVSport 2.0 780 2409 1 2409 48 48 1677 1 24 25 484 Oct. 13, 1999 404 HOFMQ33 PTA-848 pCMVSport 2.0 781 876 1 876 78 78 1678 1 24 25 266 Oct. 13, 1999 404 HOFMQ33 PTA-848 pCMVSport 2.0 782 1586 1 1586 724 1679 1 5 Oct. 13, 1999 404 HOFMQ33 PTA-848 pCMVSport 2.0 783 1011 873 1011 123 1680 1 1 2 84 Oct. 13, 1999 405 HOFMT75 PTA-848 pCMVSport 2.0 415 2131 6 2131 83 83 1312 1 20 21 410 Oct. 13, 1999 405 HOFMT75 PTA-848 pCMVSport 2.0 784 427 1 427 83 83 1681 1 20 21 115 Oct. 13, 1999 405 HOFMT75 PTA-848 pCMVSport 2.0 785 1500 1 1500 1225 1682 1 9 10 92 Oct. 13, 1999 405 HOFMT75 PTA-848 pCMVSport 2.0 786 1234 337 1234 129 129 1683 1 20 21 368 Oct. 13, 1999 406 HOFNC14 PTA-623 pCMVSport 2.0 416 2794 1 2794 79 79 1313 1 13 14 73 Sep. 02, 1999 406 HOFNC14 PTA-623 pCMVSport 2.0 787 3095 1 3095 155 155 1684 1 13 14 72 Sep. 02, 1999 407 HOFND85 PTA-1544 pCMVSport 2.0 417 2048 1 2048 167 167 1314 1 22 23 627 Mar. 21, 2000 408 HOFNY91 PTA-1544 pCMVSport 2.0 418 2406 1 2406 64 64 1315 1 14 15 82 Mar. 21, 2000 409 HOFOC33 PTA-848 pCMVSport 2.0 419 1669 1 1669 76 76 1316 1 21 22 363 Oct. 13, 1999 409 HOFOC33 PTA-848 pCMVSport 2.0 788 518 1 518 81 81 1685 1 21 22 112 Oct. 13, 1999 409 HOFOC33 PTA-848 pCMVSport 2.0 789 518 1 518 81 81 1686 1 17 18 112 Oct. 13, 1999 409 HOFOC33 PTA-848 pCMVSport 2.0 790 1670 1 1670 76 76 1687 1 21 22 139 Oct. 13, 1999 409 HOFOC33 PTA-848 pCMVSport 2.0 791 606 1 606 23 1688 1 7 Oct. 13, 1999 409 HOFOC33 PTA-848 pCMVSport 2.0 792 841 1 841 158 1689 1 6 7 14 Oct. 13, 1999 409 HOFOC33 PTA-848 pCMVSport 2.0 793 868 1 847 3 1690 1 1 2 288 Oct. 13, 1999 410 HOGCK20 209853 pCMVSport 2.0 420 2087 1 2087 57 57 1317 1 23 24 522 May 07, 1998 410 HOGCK20 209853 pCMVSport 2.0 794 2075 1 2054 53 1691 1 22 23 554 May 07, 1998 411 HOGCK63 PTA-848 pCMVSport 2.0 421 1409 310 1409 514 514 1318 1 29 30 246 Oct. 13, 1999 411 HOGCK63 PTA-848 pCMVSport 2.0 795 1697 144 1697 1455 1692 1 5 Oct. 13, 1999 412 HOGCS52 PTA-848 pCMVSport 2.0 422 2571 1 2571 25 25 1319 1 22 23 453 Oct. 13, 1999 412 HOGCS52 PTA-848 pCMVSport 2.0 796 2645 1 2586 30 30 1693 1 22 23 453 Oct. 13, 1999 412 HOGCS52 PTA-848 pCMVSport 2.0 797 1098 457 638 2 1694 1 1 2 96 Oct. 13, 1999 413 HOHBB49 203517 pCMVSport 2.0 423 3080 1 3080 148 148 1320 1 19 20 48 Dec. 10, 1998 414 HOHBC68 209568 pCMVSport 2.0 424 1837 1 1837 348 348 1321 1 30 31 128 Jan. 06, 1998 415 HOHBY12 209603 pCMVSport 2.0 425 1188 1 1188 232 232 1322 1 25 26 199 Jan. 29, 1998 416 HOHCC74 209346 pCMVSport 2.0 426 558 1 558 327 327 1323 1 20 21 48 Oct. 09, 1997 417 HOHCH55 203331 pCMVSport 2.0 427 2499 1 2499 221 221 1324 1 23 24 494 Oct. 08, 1998 417 HOHCH55 203331 pCMVSport 2.0 798 2522 1 2522 230 230 1695 1 23 24 469 Oct. 08, 1998 418 HOSDJ25 209423 Uni-ZAP XR 428 2214 985 2214 1076 1076 1325 1 18 19 40 Oct. 30, 1997 418 HOSDJ25 209423 Uni-ZAP XR 799 1258 1 1258 146 146 1696 1 18 19 40 Oct. 30, 1997 419 HOSEG51 209324 Uni-ZAP XR 429 590 48 590 232 232 1326 1 31 32 102 Oct. 02, 1997 420 HOSEQ49 209551 Uni-ZAP XR 430 1943 280 1935 544 544 1327 1 32 33 51 Dec. 12, 1997 421 HOSFD58 97957 Uni-ZAP XR 431 2527 290 1747 56 56 1328 1 30 31 624 Mar. 13, 1997 209073 May 22, 1997 421 HOSFD58 97957 Uni-ZAP XR 800 2527 288 1747 477 477 1697 1 32 33 61 Mar. 13, 1997 209073 May 22, 1997 422 HOUCQ17 209086 Uni-ZAP XR 432 4712 1 4693 508 508 1329 1 51 52 967 May 29, 1997 423 HOUDK26 209423 Uni-ZAP XR 433 1051 1 1051 214 214 1330 1 30 31 174 Oct. 30, 1997 424 HOUGG12 203071 Uni-ZAP XR 434 1895 1 1895 289 289 1331 1 20 21 314 Jul. 27, 1998 424 HOUGG12 203071 Uni-ZAP XR 801 1050 1 1050 399 399 1698 1 21 22 114 Jul. 27, 1998 424 HOUGG12 203071 Uni-ZAP XR 802 1642 35 1642 116 116 1699 1 22 23 61 Jul. 27, 1998 425 HOVCA92 209299 pSport1 435 707 1 488 181 181 1332 1 20 21 62 Sep. 25, 1997 426 HPASA81 203181 Uni-ZAP XR 436 1945 1 1945 19 19 1333 1 17 18 600 Sep. 09, 1998 426 HPASA81 203181 Uni-ZAP XR 803 1971 2 1971 14 14 1700 1 17 18 315 Sep. 09, 1998 426 HPASA81 203181 Uni-ZAP XR 804 2081 1 2081 124 124 1701 1 17 18 72 Sep. 09, 1998 427 HPBCU51 97977 pBLUESCRIPT ™ 437 599 1 599 86 86 1334 1 27 28 119 Apr. 04, 1997 SK− 209082 May 29, 1997 428 HPDDC77 209012 pBLUESCRIPT ™ 438 978 1 978 51 51 1335 1 29 30 131 Apr. 28, 1997 SK− 209089 Jun. 05, 1997 428 HPDDC77 209012 pBLUESCRIPT ™ 805 2361 455 1442 510 510 1702 1 29 30 131 Apr. 28, 1997 SK− 209089 Jun. 05, 1997 429 HPDWP28 PTA-2076 pSport1 439 528 1 528 143 143 1336 1 29 30 49 Jun. 09, 2000 429 HPDWP28 PTA-2076 pSport1 806 510 1 500 133 133 1703 1 29 30 49 Jun. 09, 2000 430 HPFCL43 209299 Uni-ZAP XR 440 665 1 665 21 21 1337 1 17 18 79 Sep. 25, 1997 431 HPFDG48 209324 Uni-ZAP XR 441 723 165 700 283 283 1338 1 18 19 47 Oct. 02, 1997 432 HPIAQ68 203517 Uni-ZAP XR 442 2466 1 2466 20 20 1339 1 22 23 62 Dec. 10, 1998 433 HPIBO15 209563 Uni-ZAP XR 443 1739 1 1739 128 128 1340 1 18 19 211 Dec. 18, 1997 433 HPIBO15 209563 Uni-ZAP XR 807 1739 1 1739 127 127 1704 1 18 19 173 Dec. 18, 1997 434 HPJBK12 PTA-855 Uni-ZAP XR 444 2648 1 2648 126 126 1341 1 18 19 48 Oct. 18, 1999 434 HPJBK12 PTA-855 Uni-ZAP XR 808 538 1 538 119 119 1705 1 18 19 48 Oct. 18, 1999 434 HPJBK12 PTA-855 Uni-ZAP XR 809 1346 1 1346 969 1706 1 10 Oct. 18, 1999 434 HPJBK12 PTA-855 Uni-ZAP XR 810 912 1 912 509 509 1707 1 4 Oct. 18, 1999 435 HPJCL22 PTA-2071 Uni-ZAP XR 445 3107 1 3107 86 86 1342 1 35 36 80 Jun. 09, 2000 435 HPJCL22 PTA-2071 Uni-ZAP XR 811 995 58 995 136 136 1708 1 35 36 80 Jun. 09, 2000 435 HPJCL22 PTA-2071 Uni-ZAP XR 812 751 183 751 232 1709 1 1 2 145 Jun. 09, 2000 436 HPJCW04 209551 Uni-ZAP XR 446 1466 1 1466 44 44 1343 1 19 20 57 Dec. 12, 1997 437 HPJEX20 PTA-872 Uni-ZAP XR 447 566 1 566 23 23 1344 1 26 27 174 Oct. 26, 1999 437 HPJEX20 PTA-872 Uni-ZAP XR 813 1823 1 1823 31 31 1710 1 23 24 115 Oct. 26, 1999 437 HPJEX20 PTA-872 Uni-ZAP XR 814 1964 1 1964 170 170 1711 1 23 24 174 Oct. 26, 1999 437 HPJEX20 PTA-872 Uni-ZAP XR 815 769 1 769 84 84 1712 1 23 24 228 Oct. 26, 1999 437 HPJEX20 PTA-872 Uni-ZAP XR 816 818 1 818 565 1713 1 1 2 84 Oct. 26, 1999 438 HPMAI22 209683 Uni-ZAP XR 448 1274 334 1274 483 483 1345 1 16 17 59 Mar. 20, 1998 439 HPMFP40 209628 Uni-ZAP XR 449 1217 1 1217 37 37 1346 1 24 25 44 Feb. 12, 1998 440 HPMGJ45 203105 Uni-ZAP XR 450 1656 1 1656 119 119 1347 1 25 26 48 Aug. 13, 1998 441 HPQAC69 97979 Lambda ZAP II 451 990 1 988 82 82 1348 1 19 20 37 Mar. 27, 1997 442 HPRBC80 209852 Uni-ZAP XR 452 2543 1245 2543 94 94 1349 1 30 31 387 May 07, 1998 442 HPRBC80 209852 Uni-ZAP XR 817 2052 275 2032 404 404 1714 1 26 27 69 May 07, 1998 443 HPRSB76 209244 pBLUESCRIPT ™ 453 741 1 741 127 127 1350 1 22 23 59 Sep. 12, 1997 444 HPVAB94 209244 Uni-ZAP XR 454 819 1 819 80 80 1351 1 25 26 44 Sep. 12, 1997 445 HPWAY46 PTA-843 Uni-ZAP XR 455 1414 1 1414 468 468 1352 1 30 31 52 Oct. 13, 1999 445 HPWAY46 PTA-843 Uni-ZAP XR 818 891 1 891 474 474 1715 1 30 31 52 Oct. 13, 1999 445 HPWAY46 PTA-843 Uni-ZAP XR 819 501 120 501 178 1716 1 1 2 86 Oct. 13, 1999 446 HPWAZ95 209007 Uni-ZAP XR 456 323 1 323 88 88 1353 1 27 28 78 Apr. 28, 1997 209083 May 29, 1997 447 HPWDJ42 209852 Uni-ZAP XR 457 1340 1 1340 149 149 1354 1 18 19 54 May 07, 1998 447 HPWDJ42 209852 Uni-ZAP XR 820 1340 1 1340 149 149 1717 1 21 22 54 May 07, 1998 447 HPWDJ42 209852 Uni-ZAP XR 821 813 1 813 161 161 1718 1 18 19 47 May 07, 1998 448 HPZAB47 209511 pBLUESCRIPT ™ 458 1676 1 1676 34 34 1355 1 18 19 47 Dec. 03, 1997 449 HRAAB15 209651 pCMVSport 3.0 459 1747 1 1747 35 35 1356 1 14 15 159 Mar. 04, 1998 450 HRABA80 209889 pCMVSport 3.0 460 1251 1 1251 144 144 1357 1 27 28 102 May 22, 1998 450 HRABA80 209889 pCMVSport 3.0 822 1237 1 1237 130 130 1719 1 27 28 102 May 22, 1998 451 HRACD15 209852 pCMVSport 3.0 461 1539 24 1539 252 252 1358 1 40 41 53 May 07, 1998 451 HRACD15 209852 pCMVSport 3.0 823 1681 24 1453 252 252 1720 1 40 41 53 May 07, 1998 452 HRACD80 209889 pCMVSport 3.0 462 1941 1 1941 196 196 1359 1 16 17 575 May 22, 1998 452 HRACD80 209889 pCMVSport 3.0 824 1934 1 1934 191 191 1721 1 16 17 575 May 22, 1998 452 HRACD80 209889 pCMVSport 3.0 825 1958 1 1958 191 191 1722 1 16 17 146 May 22, 1998 453 HRDDV47 209628 Uni-ZAP XR 463 1510 1 1510 146 146 1360 1 30 31 276 Feb. 12, 1998 454 HRDFD27 209423 Uni-ZAP XR 464 805 1 805 82 82 1361 1 35 36 83 Oct. 30, 1997 455 HRTAE58 209241 pBLUESCRIPT ™ 465 600 1 600 244 244 1362 1 18 19 58 Sep. 12, 1997 SK− 456 HSATR82 209299 Uni-ZAP XR 466 777 1 777 74 74 1363 1 15 16 41 Sep. 25, 1997 457 HSAUK57 209148 Uni-ZAP XR 467 1037 1 1037 322 322 1364 1 26 27 83 Jul. 17, 1997 457 HSAUK57 209148 Uni-ZAP XR 826 1070 1 1070 327 327 1723 1 26 27 48 Jul. 17, 1997 458 HSAUL82 209148 Uni-ZAP XR 468 727 1 727 140 140 1365 1 25 26 49 Jul. 17, 1997 459 HSAVD46 209124 Uni-ZAP XR 469 773 2 767 155 155 1366 1 20 21 58 Jun. 19, 1997 460 HSAVH65 209651 Uni-ZAP XR 470 600 1 600 104 104 1367 1 21 22 100 Mar. 04, 1998 461 HSAVK10 209368 Uni-ZAP XR 471 1242 1 1242 131 131 1368 1 32 33 40 Oct. 16, 1997 462 HSAWZ41 209463 Uni-ZAP XR 472 1388 1 1388 98 98 1369 1 24 25 57 Nov. 14, 1997 463 HSAXA83 209324 Uni-ZAP XR 473 649 1 649 92 92 1370 1 22 23 74 Oct. 02, 1997 464 HSAYM40 209139 Uni-ZAP XR 474 433 1 433 190 190 1371 1 19 20 63 Jul. 03, 1997 465 HSDAJ46 209746 Uni-ZAP XR 475 1537 92 1537 299 299 1372 1 18 19 262 Apr. 07, 1998 466 HSDEK49 209603 Uni-ZAP XR 476 1782 1 1782 60 60 1373 1 19 20 399 Jan. 29, 1998 466 HSDEK49 209603 Uni-ZAP XR 827 1590 96 1590 126 126 1724 1 21 22 305 Jan. 29, 1998 467 HSDER95 209683 Uni-ZAP XR 477 574 1 574 72 72 1374 1 25 26 71 Mar. 20, 1998 468 HSDEZ20 209852 Uni-ZAP XR 478 795 1 795 58 58 1375 1 41 42 122 May 07, 1998 468 HSDEZ20 209852 Uni-ZAP XR 828 1540 1 1540 66 66 1725 1 41 42 97 May 07, 1998 469 HSDJA15 203081 Uni-ZAP XR 479 1443 1 1443 247 247 1376 1 20 21 152 Jul. 30, 1998 470 HSDSB09 209145 pBLUESCRIPT ™ 480 809 1 809 16 16 1377 1 17 18 135 Jul. 17, 1997 470 HSDSB09 209145 pBLUESCRIPT ™ 829 819 1 819 22 22 1726 1 17 18 121 Jul. 17, 1997 471 HSDSE75 209324 pBLUESCRIPT ™ 481 1151 1 1151 160 160 1378 1 18 19 181 Oct. 02, 1997 472 HSFAM31 209346 Uni-ZAP XR 482 868 1 868 44 44 1379 1 9 Oct. 09, 1997 473 HSHAX21 209853 Uni-ZAP XR 483 1986 1 1986 177 177 1380 1 13 14 72 May 07, 1998 474 HSIAS17 209226 Uni-ZAP XR 484 1781 1 1781 431 431 1381 1 22 23 257 Aug. 28, 1997 474 HSIAS17 209226 Uni-ZAP XR 830 1448 1 1224 108 108 1727 1 23 24 218 Aug. 28, 1997 475 HSIDX71 PTA-843 Uni-ZAP XR 485 2118 1 2118 200 200 1382 1 41 42 59 Oct. 13, 1999 475 HSIDX71 PTA-843 Uni-ZAP XR 831 1868 1 1868 200 200 1728 1 41 42 59 Oct. 13, 1999 476 HSKDA27 PTA-322 Uni-ZAP XR 486 4412 1 4412 786 786 1383 1 24 25 950 Jul. 09, 1999 476 HSKDA27 PTA-322 Uni-ZAP XR 832 1792 134 1792 127 127 1729 1 21 22 509 Jul. 09, 1999 476 HSKDA27 PTA-322 Uni-ZAP XR 833 1673 1 1673 12 12 1730 1 21 22 554 Jul. 09, 1999 477 HSKHZ81 209346 pBLUESCRIPT ™ 487 969 1 969 64 64 1384 1 27 28 247 Oct. 09, 1997 477 HSKHZ81 209346 pBLUESCRIPT ™ 834 988 1 967 57 57 1731 1 27 28 247 Oct. 09, 1997 478 HSLCQ82 209551 Uni-ZAP XR 488 1476 1 1476 226 226 1385 1 28 29 84 Dec. 12, 1997 478 HSLCQ82 209551 Uni-ZAP XR 835 1501 1 1501 233 233 1732 1 22 23 57 Dec. 12, 1997 479 HSLJG37 PTA-855 Uni-ZAP XR 489 2126 1 2126 114 114 1386 1 16 17 42 Oct. 18, 1999 479 HSLJG37 PTA-855 Uni-ZAP XR 836 1083 1 1083 206 206 1733 1 16 17 42 Oct. 18, 1999 479 HSLJG37 PTA-855 Uni-ZAP XR 837 1904 1 1904 1331 1734 1 6 Oct. 18, 1999 480 HSNAB12 209300 Uni-ZAP XR 490 630 1 630 151 151 1387 1 27 28 71 Sep. 25, 1997 481 HSODE04 PTA-855 Uni-ZAP XR 491 1370 1 1370 202 202 1388 1 20 21 41 Oct. 18, 1999 481 HSODE04 PTA-855 Uni-ZAP XR 838 1937 1 1937 300 300 1735 1 20 21 41 Oct. 18, 1999 482 HSPBF70 203105 pSport1 492 1397 288 1397 429 429 1389 1 19 20 97 Aug. 13, 1998 483 HSQCM10 209641 Uni-ZAP XR 493 657 1 654 130 130 1390 1 19 20 62 Feb. 25, 1998 484 HSSAJ29 209626 Uni-ZAP XR 494 1044 1 1044 103 103 1391 1 25 26 47 Feb. 12, 1998 485 HSSDX51 209683 Uni-ZAP XR 495 1143 1 1143 133 133 1392 1 20 21 50 Mar. 20, 1998 486 HSSFT08 209551 Uni-ZAP XR 496 791 1 791 125 125 1393 1 34 35 58 Dec. 12, 1997 487 HSSGD52 PTA-1543 Uni-ZAP XR 497 2425 1 2425 344 344 1394 1 32 33 606 Mar. 21, 2000 487 HSSGD52 PTA-1543 Uni-ZAP XR 839 2460 105 2460 338 338 1736 1 27 28 606 Mar. 21, 2000 488 HSSJC35 209853 Uni-ZAP XR 498 1174 1 1174 62 62 1395 1 28 29 295 May 07, 1998 488 HSSJC35 209853 Uni-ZAP XR 840 1163 1 1163 55 55 1737 1 30 31 295 May 07, 1998 488 HSSJC35 209853 Uni-ZAP XR 841 1183 1 1183 66 66 1738 1 30 31 37 May 07, 1998 489 HSTBJ86 203027 Uni-ZAP XR 499 1766 1 1766 120 120 1396 1 24 25 83 Jun. 26, 1998 490 HSUBW09 209007 Uni-ZAP XR 500 1021 1 1021 153 153 1397 1 31 32 56 Apr. 28, 1997 209083 May 29, 1997 491 HSVAM10 209244 Uni-ZAP XR 501 433 1 433 46 46 1398 1 27 28 51 Sep. 12, 1997 492 HSVBU91 209603 Uni-ZAP XR 502 727 1 727 256 256 1399 1 18 19 90 Jan. 29, 1998 493 HSXCG83 203570 Uni-ZAP XR 503 2112 233 1573 101 101 1400 1 45 46 267 Jan. 11, 1999 493 HSXCG83 203570 Uni-ZAP XR 842 1938 58 1399 211 211 1739 1 22 23 172 Jan. 11, 1999 494 HSXEC75 209641 Uni-ZAP XR 504 1112 1 1112 295 295 1401 1 33 34 45 Feb. 25, 1998 495 HSXEQ06 PTA-847 Uni-ZAP XR 505 1598 1 1598 123 123 1402 1 24 25 60 Oct. 13, 1999 495 HSXEQ06 PTA-847 Uni-ZAP XR 843 768 21 768 136 136 1740 1 24 25 60 Oct. 13, 1999 495 HSXEQ06 PTA-847 Uni-ZAP XR 844 1392 1 1392 1271 1741 1 9 10 17 Oct. 13, 1999 496 HSYAV50 PTA-1544 pCMVSport 3.0 506 2801 1 2801 155 155 1403 1 23 24 672 Mar. 21, 2000 497 HSYAV66 209746 pCMVSport 3.0 507 1407 1 1407 186 186 1404 1 28 29 69 Apr. 07, 1998 498 HSYAZ50 PTA-849 pCMVSport 3.0 508 1097 1 1097 131 131 1405 1 18 19 56 Oct. 13, 1999 498 HSYAZ50 PTA-849 pCMVSport 3.0 845 768 226 768 345 345 1742 1 18 19 56 Oct. 13, 1999 498 HSYAZ50 PTA-849 pCMVSport 3.0 846 2087 770 875 723 1743 1 1 2 106 Oct. 13, 1999 498 HSYAZ50 PTA-849 pCMVSport 3.0 847 2096 1767 2050 2 1744 1 1 2 279 Oct. 13, 1999 499 HSYAZ63 PTA-163 pCMVSport 3.0 509 3466 1655 3347 448 448 1406 1 30 31 434 Jun. 01, 1999 499 HSYAZ63 PTA-163 pCMVSport 3.0 848 1707 1 1707 215 215 1745 1 21 22 40 Jun. 01, 1999 500 HSYBG37 209463 pCMVSport 3.0 510 1238 1 1238 47 47 1407 1 24 25 305 Nov. 14, 1997 500 HSYBG37 209463 pCMVSport 3.0 849 1239 1 1239 48 48 1746 1 24 25 305 Nov. 14, 1997 501 HSZAF47 209124 Uni-ZAP XR 511 1304 1 1304 106 106 1408 1 16 17 289 Jun. 19, 1997 501 HSZAF47 209124 Uni-ZAP XR 850 1333 2 1333 107 107 1747 1 18 19 127 Jun. 19, 1997 502 HT3SF53 PTA-499 Uni-ZAP XR 512 1926 1 1926 184 184 1409 1 27 28 68 Aug. 11, 1999 503 HT5GJ57 209889 Uni-ZAP XR 513 1773 1 1773 105 105 1410 1 25 26 243 May 22, 1998 503 HT5GJ57 209889 Uni-ZAP XR 851 1797 92 1797 122 122 1748 1 25 26 190 May 22, 1998 504 HTADX17 209124 Uni-ZAP XR 514 1147 0 1148 92 92 1411 1 23 24 142 Jun. 19, 1997 504 HTADX17 209124 Uni-ZAP XR 852 1140 22 1140 84 84 1749 1 19 20 142 Jun. 19, 1997 505 HTDAF28 97974 pSport1 515 912 1 912 38 38 1412 1 22 23 87 Apr. 04, 1997 209080 May 29, 1997 506 HTEAF65 PTA-322 Uni-ZAP XR 516 563 1 563 135 135 1413 1 19 20 75 Jul. 09, 1999 507 HTEBI28 209177 Uni-ZAP XR 517 413 1 413 43 43 1414 1 20 21 67 Jul. 24, 1997 508 HTEDF80 209511 Uni-ZAP XR 518 1306 1 1306 696 696 1415 1 21 22 126 Dec. 03, 1997 509 HTEDY42 209241 Uni-ZAP XR 519 754 1 754 19 19 1416 1 23 24 233 Sep. 12, 1997 509 HTEDY42 209241 Uni-ZAP XR 853 810 1 810 19 19 1750 1 23 24 77 Sep. 12, 1997 510 HTEFU65 209324 Uni-ZAP XR 520 1028 1 1028 231 231 1417 1 24 25 46 Oct. 02, 1997 511 HTEGI42 PTA-842 Uni-ZAP XR 521 978 1 978 26 26 1418 1 19 20 257 Oct. 13, 1999 511 HTEGI42 PTA-842 Uni-ZAP XR 854 1092 1 1092 145 145 1751 1 19 20 257 Oct. 13, 1999 511 HTEGI42 PTA-842 Uni-ZAP XR 855 284 1 133 1 1752 1 1 2 94 Oct. 13, 1999 511 HTEGI42 PTA-842 Uni-ZAP XR 856 1494 754 937 1081 1753 1 1 2 82 Oct. 13, 1999 511 HTEGI42 PTA-842 Uni-ZAP XR 857 1014 1 806 670 1754 1 1 2 60 Oct. 13, 1999 512 HTEHR24 209224 Uni-ZAP XR 522 1075 50 1075 84 84 1419 1 29 30 163 Aug. 28, 1997 512 HTEHR24 209224 Uni-ZAP XR 858 1038 1 1038 41 41 1755 1 28 29 124 Aug. 28, 1997 513 HTEHU31 209568 Uni-ZAP XR 523 1113 1 1113 121 121 1420 1 25 26 312 Jan. 06, 1998 514 HTEHU93 209090 Uni-ZAP XR 524 738 1 738 188 188 1421 1 24 25 142 Jun. 05, 1997 514 HTEHU93 209090 Uni-ZAP XR 859 745 1 745 187 187 1756 1 24 25 113 Jun. 05, 1997 515 HTEIP36 209244 Uni-ZAP XR 525 752 1 752 22 22 1422 1 19 20 58 Sep. 12, 1997 516 HTEIV80 209511 Uni-ZAP XR 526 1748 1 1748 203 203 1423 1 14 15 47 Dec. 03, 1997 517 HTEJN13 97958 Uni-ZAP XR 527 1094 1 1094 156 156 1424 1 15 16 208 Mar. 13, 1997 209072 May 22, 1997 517 HTEJN13 97958 Uni-ZAP XR 860 1147 1 1147 163 163 1757 1 15 16 159 Mar. 13, 1997 209072 May 22, 1997 517 HTEJN13 97958 Uni-ZAP XR 861 1134 1 1134 155 155 1758 1 19 20 71 Mar. 13, 1997 209072 May 22, 1997 518 HTELM16 203648 Uni-ZAP XR 528 531 1 531 121 121 1425 1 21 22 84 Feb. 09, 1999 519 HTEPG70 203570 Uni-ZAP XR 529 813 1 813 365 365 1426 1 27 28 89 Jan. 11, 1999 520 HTGAU75 209563 Uni-ZAP XR 530 1713 1 1713 149 149 1427 1 33 34 142 Dec. 18, 1997 521 HTGEP89 97977 Uni-ZAP XR 531 703 1 703 285 285 1428 1 29 30 94 Apr. 04, 1997 209082 May 29, 1997 522 HTHBG43 PTA-843 Uni-ZAP XR 532 848 1 848 47 47 1429 1 39 Oct. 13, 1999 522 HTHBG43 PTA-843 Uni-ZAP XR 862 632 103 632 149 149 1759 1 39 Oct. 13, 1999 523 HTHCA18 PTA-844 Uni-ZAP XR 533 1818 1 1818 231 231 1430 1 15 16 38 Oct. 13, 1999 523 HTHCA18 PTA-844 Uni-ZAP XR 863 2036 1 2036 224 224 1760 1 15 16 38 Oct. 13, 1999 524 HTHDJ94 209746 Uni-ZAP XR 534 1632 20 1632 66 66 1431 1 26 27 292 Apr. 07, 1998 525 HTHDS25 203071 Uni-ZAP XR 535 1061 1 1061 70 70 1432 1 15 16 90 Jul. 27, 1998 526 HTJMA95 209853 pCMVSport 2.0 536 1650 198 1569 527 527 1433 1 22 23 181 May 07, 1998 527 HTJML75 PTA-868 pCMVSport 2.0 537 2762 1 2762 30 30 1434 1 1 2 822 Oct. 26, 1999 527 HTJML75 PTA-868 pCMVSport 2.0 864 2694 21 2694 335 1761 1 20 21 64 Oct. 26, 1999 528 HTLBE23 PTA-842 Uni-ZAP XR 538 1216 1 1216 129 129 1435 1 17 18 45 Oct. 13, 1999 528 HTLBE23 PTA-842 Uni-ZAP XR 865 810 286 810 205 1762 1 5 Oct. 13, 1999 529 HTLFE42 209138 Uni-ZAP XR 539 712 1 712 116 116 1436 1 22 23 77 Jul. 03, 1997 530 HTLFE57 PTA-1543 Uni-ZAP XR 540 2248 1 2248 124 124 1437 1 17 18 188 Mar. 21, 2000 530 HTLFE57 PTA-1543 Uni-ZAP XR 866 2298 1157 2214 189 189 1763 1 18 19 170 Mar. 21, 2000 530 HTLFE57 PTA-1543 Uni-ZAP XR 867 928 1 928 110 110 1764 1 18 19 170 Mar. 21, 2000 531 HTLGE31 PTA-2081 Uni-ZAP XR 541 534 1 534 51 51 1438 1 17 18 86 Jun. 09, 2000 532 HTLHY14 203648 Uni-ZAP XR 542 1032 1 1032 36 36 1439 1 17 18 246 Feb. 09, 1999 533 HTLIT32 203570 Uni-ZAP XR 543 1074 164 897 288 288 1440 1 26 27 246 Jan. 11, 1999 534 HTLIV19 PTA-2081 Uni-ZAP XR 544 978 1 978 110 110 1441 1 33 34 84 Jun. 09, 2000 535 HTNBO91 209241 pBLUESCRIPT ™ 545 300 1 300 7 7 1442 1 26 27 40 Sep. 12, 1997 SK− 536 HTOAK16 209368 Uni-ZAP XR 546 1466 1 1466 87 87 1443 1 18 19 110 Oct. 16, 1997 537 HTODK73 209244 Uni-ZAP XR 547 1019 4 1019 43 43 1444 1 23 24 59 Sep. 12, 1997 538 HTODO72 209299 Uni-ZAP XR 548 973 1 973 183 183 1445 1 16 17 24 Sep. 25, 1997 539 HTOGR42 209603 Uni-ZAP XR 549 1430 1 1430 14 14 1446 1 18 19 56 Jan. 29, 1998 539 HTOGR42 209603 Uni-ZAP XR 868 1433 1 1433 13 13 1765 1 18 19 60 Jan. 29, 1998 540 HTOHD42 203081 Uni-ZAP XR 550 946 1 946 155 155 1447 1 24 25 190 Jul. 30, 1998 541 HTOHM15 PTA-843 Uni-ZAP XR 551 1949 1 1949 30 30 1448 1 20 21 61 Oct. 13, 1999 541 HTOHM15 PTA-843 Uni-ZAP XR 869 408 1 408 23 23 1766 1 20 21 61 Oct. 13, 1999 541 HTOHM15 PTA-843 Uni-ZAP XR 870 1299 982 1274 71 1767 1 1 2 322 Oct. 13, 1999 541 HTOHM15 PTA-843 Uni-ZAP XR 871 1669 1 1622 1555 1768 1 9 10 13 Oct. 13, 1999 542 HTOHT18 209745 Uni-ZAP XR 552 1499 267 1499 433 433 1449 1 24 25 53 Apr. 07, 1998 543 HTOIZ02 PTA-843 Uni-ZAP XR 553 549 1 549 243 243 1450 1 16 17 50 Oct. 13, 1999 543 HTOIZ02 PTA-843 Uni-ZAP XR 872 1369 746 1345 2 1769 1 1 2 240 Oct. 13, 1999 544 HTOJA73 203105 Uni-ZAP XR 554 1294 1 1294 100 100 1451 1 21 22 41 Aug. 13, 1998 545 HTOJK60 209324 Uni-ZAP XR 555 904 1 904 217 217 1452 1 18 19 32 Oct. 02, 1997 546 HTPBW79 209511 Uni-ZAP XR 556 1374 1 1374 178 178 1453 1 22 23 362 Dec. 03, 1997 546 HTPBW79 209511 Uni-ZAP XR 873 1515 118 1507 302 302 1770 1 24 25 362 Dec. 03, 1997 546 HTPBW79 209511 Uni-ZAP XR 874 1404 1 1404 92 92 1771 1 22 23 415 Dec. 03, 1997 547 HTSEW17 209138 pBLUESCRIPT ™ 557 652 1 652 170 170 1454 1 34 35 37 Jul. 03, 1997 548 HTTBI76 209641 Uni-ZAP XR 558 1711 1 1711 133 133 1455 1 22 23 133 Feb. 25, 1998 549 HTTDB46 203484 Uni-ZAP XR 559 3059 1 3059 55 55 1456 1 17 18 318 Nov. 17, 1998 549 HTTDB46 203484 Uni-ZAP XR 875 2008 215 2008 153 153 1772 1 17 18 461 Nov. 17, 1998 550 HTWCT03 209086 pSport1 560 1963 1 1963 334 334 1457 1 26 27 101 May 29, 1997 551 HTWDF76 209852 pSport1 561 963 1 963 316 316 1458 1 24 25 85 May 07, 1998 552 HTWJK32 209852 Lambda ZAP II 562 911 211 911 376 376 1459 1 20 21 51 May 07, 1998 553 HTWKE60 209651 Lambda ZAP II 563 407 1 407 185 185 1460 1 25 26 44 Mar. 04, 1998 554 HTXCV12 209423 Uni-ZAP XR 564 1134 1 1134 175 175 1461 1 27 28 102 Oct. 30, 1997 554 HTXCV12 209423 Uni-ZAP XR 876 1162 1 1162 183 183 1773 1 27 28 91 Oct. 30, 1997 555 HTXDW56 209746 Uni-ZAP XR 565 1583 1 1583 217 217 1462 1 21 22 201 Apr. 07, 1998 556 HTXFL30 209603 Uni-ZAP XR 566 1991 1 1991 30 30 1463 1 39 40 102 Jan. 29, 1998 557 HTXKP61 203364 Uni-ZAP XR 567 1209 1 1209 169 169 1464 1 33 34 42 Oct. 19, 1998 558 HUDBZ89 209407 ZAP Express 568 2135 1 2135 1085 1085 1465 1 17 18 73 Oct. 23, 1997 558 HUDBZ89 209407 ZAP Express 877 1265 1 1265 197 197 1774 1 17 18 54 Oct. 23, 1997 559 HUFBY15 PTA-1543 pSport1 569 1193 1 1193 49 49 1466 1 26 27 159 Mar. 21, 2000 559 HUFBY15 PTA-1543 pSport1 878 1012 1 1012 74 74 1775 1 26 27 145 Mar. 21, 2000 560 HUFEF62 209852 pSport1 570 518 1 518 190 190 1467 1 28 29 68 May 07, 1998 560 HUFEF62 209852 pSport1 879 539 1 539 182 182 1776 1 28 29 68 May 07, 1998 561 HUKAH51 209568 Lambda ZAP II 571 853 1 853 286 286 1468 1 20 21 151 Jan. 06, 1998 561 HUKAH51 209568 Lambda ZAP II 880 754 1 754 144 144 1777 1 22 23 142 Jan. 06, 1998 561 HUKAH51 209568 Lambda ZAP II 881 667 1 667 55 55 1778 1 22 23 119 Jan. 06, 1998 562 HUKBT29 209746 Lambda ZAP II 572 1757 56 1757 74 74 1469 1 19 20 506 Apr. 07, 1998 563 HUSAT94 209580 Lambda ZAP II 573 2234 269 2234 302 302 1470 1 28 29 45 Jan. 14, 1998 564 HUSBA88 PTA-623 Lambda ZAP II 574 2733 27 2733 270 270 1471 1 15 16 615 Sep. 02, 1999 565 HUSIG64 209423 pSport1 575 1010 1 1010 9 9 1472 1 21 22 334 Oct. 30, 1997 566 HUSXS50 209651 pSport1 576 2561 1 2561 280 280 1473 1 19 20 522 Mar. 04, 1998 566 HUSXS50 209651 pSport1 882 2025 1098 1997 281 281 1779 1 30 31 462 Mar. 04, 1998 566 HUSXS50 209651 pSport1 883 1020 1 1020 179 179 1780 1 23 24 174 Mar. 04, 1998 567 HWAAD63 203570 pCMVSport 3.0 577 3308 1 3308 322 322 1474 1 30 31 168 Jan. 11, 1999 567 HWAAD63 203570 pCMVSport 3.0 884 3306 1 3306 322 322 1781 1 30 31 53 Jan. 11, 1999 567 HWAAD63 203570 pCMVSport 3.0 885 2194 1 2194 312 312 1782 1 30 31 169 Jan. 11, 1999 568 HWABA81 209463 pCMVSport 3.0 578 866 1 866 57 57 1475 1 21 22 48 Nov. 14, 1997 569 HWABY10 203071 pCMVSport 3.0 579 2950 78 2914 263 263 1476 1 22 23 168 Jul. 27, 1998 570 HWADJ89 PTA-1543 pCMVSport 3.0 580 1769 529 1769 581 581 1477 1 1 2 43 Mar. 21, 2000 571 HWBAO62 209603 pCMVSport 3.0 581 1903 1 1903 52 52 1478 1 30 31 212 Jan. 29, 1998 571 HWBAO62 209603 pCMVSport 3.0 886 1940 1 1940 81 81 1783 1 30 31 101 Jan. 29, 1998 572 HWBAR14 PTA-867 pCMVSport 3.0 582 3878 1 3878 152 152 1479 1 48 49 371 Oct. 26, 1999 572 HWBAR14 PTA-867 pCMVSport 3.0 887 432 1 432 287 287 1784 1 33 34 48 Oct. 26, 1999 572 HWBAR14 PTA-867 pCMVSport 3.0 888 794 1 794 204 1785 1 10 11 12 Oct. 26, 1999 572 HWBAR14 PTA-867 pCMVSport 3.0 889 1019 1 1019 492 1786 1 1 2 129 Oct. 26, 1999 573 HWBAR88 PTA-867 pCMVSport 3.0 583 1051 1 1051 156 156 1480 1 18 19 75 Oct. 26, 1999 574 HWBCB89 PTA-499 pCMVSport 3.0 584 1317 3 1317 37 37 1481 1 19 20 187 Aug. 11, 1999 574 HWBCB89 PTA-499 pCMVSport 3.0 890 1315 1 1315 35 35 1787 1 19 20 187 Aug. 11, 1999 575 HWBCP79 209641 pCMVSport 3.0 585 1138 1 1138 243 243 1482 1 21 22 105 Feb. 25, 1998 575 HWBCP79 209641 pCMVSport 3.0 891 1138 1 1138 233 233 1788 1 21 22 105 Feb. 25, 1998 576 HWBDP28 209641 pCMVSport 3.0 586 1841 1 1841 1342 1342 1483 1 25 26 67 Feb. 25, 1998 576 HWBDP28 209641 pCMVSport 3.0 892 314 1 314 132 132 1789 1 25 26 61 Feb. 25, 1998 577 HWBEM18 PTA-868 pCMVSport 3.0 587 6729 1 6729 75 75 1484 1 25 26 1887 Oct. 26, 1999 577 HWBEM18 PTA-868 pCMVSport 3.0 893 3599 1 3599 65 65 1790 1 25 26 886 Oct. 26, 1999 577 HWBEM18 PTA-868 pCMVSport 3.0 894 2924 1 2496 1 1791 1 1 2 498 Oct. 26, 1999 578 HWBFE57 PTA-868 pCMVSport 3.0 588 1133 36 1133 227 227 1485 1 36 37 302 Oct. 26, 1999 578 HWBFE57 PTA-868 pCMVSport 3.0 895 5811 3302 5811 3300 1792 1 16 17 37 Oct. 26, 1999 578 HWBFE57 PTA-868 pCMVSport 3.0 896 1012 1 1012 622 1793 1 10 11 16 Oct. 26, 1999 579 HWDAC39 209641 pCMVSport 3.0 589 753 1 753 96 96 1486 1 20 21 110 Feb. 25, 1998 579 HWDAC39 209641 pCMVSport 3.0 897 734 1 734 85 85 1794 1 20 21 117 Feb. 25, 1998 580 HWDAH38 PTA-868 pCMVSport 3.0 590 1604 1 1604 255 255 1487 1 20 21 40 Oct. 26, 1999 580 HWDAH38 PTA-868 pCMVSport 3.0 898 796 1 796 319 319 1795 1 20 21 40 Oct. 26, 1999 581 HWHGP71 203858 pCMVSport 3.0 591 1021 1 1021 389 389 1488 1 51 52 211 Mar. 18, 1999 581 HWHGP71 203858 pCMVSport 3.0 899 1037 1 1037 394 394 1796 1 18 19 77 Mar. 18, 1999 582 HWHGQ49 209641 pCMVSport 3.0 592 985 1 985 511 511 1489 1 17 18 90 Feb. 25, 1998 582 HWHGQ49 209641 pCMVSport 3.0 900 1410 33 1410 306 306 1797 1 22 23 150 Feb. 25, 1998 583 HWHGU54 209782 pCMVSport 3.0 593 1445 1 1445 145 145 1490 1 19 20 414 Apr. 20, 1998 584 HWHGZ51 PTA-499 pCMVSport 3.0 594 1699 1 1699 33 33 1491 1 30 31 346 Aug. 11, 1999 585 HWHHL34 203181 pCMVSport 3.0 595 1529 95 1529 131 131 1492 1 30 31 188 Sep. 09, 1998 585 HWHHL34 203181 pCMVSport 3.0 901 1796 1 1796 209 209 1798 1 31 32 102 Sep. 09, 1998 585 HWHHL34 203181 pCMVSport 3.0 902 2136 1 2136 101 101 1799 1 30 31 188 Sep. 09, 1998 586 HWHQS55 203027 pCMVSport 3.0 596 3282 1 3282 169 169 1493 1 26 27 742 Jun. 26, 1998 587 HWLEV32 PTA-884 pSport1 597 1218 1 1218 39 39 1494 1 18 19 45 Oct. 28, 1999 587 HWLEV32 PTA-884 pSport1 903 1203 1 1203 29 29 1800 1 18 19 45 Oct. 28, 1999 587 HWLEV32 PTA-884 pSport1 904 1144 528 596 3 1801 1 1 2 136 Oct. 28, 1999 587 HWLEV32 PTA-884 pSport1 905 1120 791 851 1 1802 1 1 2 141 Oct. 28, 1999 588 HWLIH65 203081 pSport1 598 831 1 831 129 129 1495 1 18 19 165 Jul. 30, 1998 589 HYAAJ71 203517 pCMVSport 3.0 599 3337 1 3337 190 190 1496 1 31 32 62 Dec. 10, 1998 590 HYBAR01 209580 Uni-ZAP XR 600 1440 1 1440 157 157 1497 1 26 27 46 Jan. 14, 1998 591 HYBBE75 203570 Uni-ZAP XR 601 838 1 838 319 319 1498 1 25 26 41 Jan. 11, 1999 592 HAPSA79 PTA-1543 Uni-ZAP XR 602 4386 1 4386 468 468 1499 1 30 31 310 Mar. 21, 2000 592 HAPSA79 PTA-1543 Uni-ZAP XR 906 4385 1 4385 468 468 1803 1 30 31 310 Mar. 21, 2000 592 HAPSA79 PTA-1543 Uni-ZAP XR 907 4386 1 4386 468 468 1804 1 30 31 310 Mar. 21, 2000

Table 1B (Comprised of Tables 1B.1 and 1B.2)

The first column in Table 1B.1 and Table 1B.2 provides the gene number in the application corresponding to the clone identifier. The second column in Table 1B.1 and Table 1B.2 provides a unique “Clone ID:” for the cDNA clone related to each contig sequence disclosed in Table 1B.1 and Table 1B.2. This clone ID references the cDNA clone which contains at least the 5′ most sequence of the assembled contig and at least a portion of SEQ ID NO:X as determined by directly sequencing the referenced clone. The referenced clone may have more sequence than described in the sequence listing or the clone may have less. In the vast majority of cases, however, the clone is believed to encode a full-length polypeptide. In the case where a clone is not full-length, a full-length cDNA can be obtained by methods described elsewhere herein. The third column in Table 1B.1 and Table 1B.2 provides a unique “Contig ID” identification for each contig sequence. The fourth column in Table 1B.1 and Table 1B.2 provides the “SEQ ID NO:” identifier for each of the contig polynucleotide sequences disclosed in Table 1B.

Table 1B.1

The fifth column in Table 1B.1, “ORF (From-To)”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence “SEQ ID NO:X” that delineate the preferred open reading frame (ORF) shown in the sequence listing and referenced in Table 1B.1, column 6, as SEQ ID NO:Y. Where the nucleotide position number “To” is lower than the nucleotide position number “From”, the preferred ORF is the reverse complement of the referenced polynucleotide sequence. The sixth column in Table 1B.1 provides the corresponding SEQ ID NO:Y for the polypeptide sequence encoded by the preferred ORF delineated in column 5. In one embodiment, the invention provides an amino acid sequence comprising, or alternatively consisting of, a polypeptide encoded by the portion of SEQ ID NO:X delineated by “ORF (From-To)”. Also provided are polynucleotides encoding such amino acid sequences and the complementary strand thereto. Column 7 in Table 1B.1 lists residues comprising epitopes contained in the polypeptides encoded by the preferred ORF (SEQ ID NO:Y), as predicted using the algorithm of Jameson and Wolf, (1988) Comp. Appl. Biosci. 4:181-186. The Jameson-Wolf antigenic analysis was performed using the computer program PROTEAN (Version 3.11 for the Power MacIntosh, DNASTAR, Inc., 1228 South Park Street Madison, Wis.). In specific embodiments, polypeptides of the invention comprise, or alternatively consist of, at least one, two, three, four, five or more of the predicted epitopes as described in Table 1B. It will be appreciated that depending on the analytical criteria used to predict antigenic determinants, the exact address of the determinant may vary slightly.

Column 8 in Table 1B.1 provides a chromosomal map location for certain polynucleotides of the invention. Chromosomal location was determined by finding exact matches to EST and cDNA sequences contained in the NCBI (National Center for Biotechnology Information) UniGene database. Each sequence in the UniGene database is assigned to a “cluster”; all of the ESTs, cDNAs, and STSs in a cluster are believed to be derived from a single gene. Chromosomal mapping data is often available for one or more sequence(s) in a UniGene cluster; this data (if consistent) is then applied to the cluster as a whole. Thus, it is possible to infer the chromosomal location of a new polynucleotide sequence by determining its identity with a mapped UniGene cluster.

A modified version of the computer program BLASTN (Altshul, et al., J. Mol. Biol. 215:403-410 (1990), and Gish, and States, Nat. Genet. 3:266-272) (1993) was used to search the UniGene database for EST or cDNA sequences that contain exact or near-exact matches to a polynucleotide sequence of the invention (the ‘Query’). A sequence from the UniGene database (the ‘Subject’) was said to be an exact match if it contained a segment of 50 nucleotides in length such that 48 of those nucleotides were in the same order as found in the Query sequence. If all of the matches that met this criteria were in the same UniGene cluster, and mapping data was available for this cluster, it is indicated in Table 1B under the heading “Cytologic Band”. Where a cluster had been further localized to a distinct cytologic band, that band is disclosed; where no banding information was available, but the gene had been localized to a single chromosome, the chromosome is disclosed.

Once a presumptive chromosomal location was determined for a polynucleotide of the invention, an associated disease locus was identified by comparison with a database of diseases which have been experimentally associated with genetic loci. The database used was the Morbid Map, derived from OMIM™ and National Center for Biotechnology Information, National Library of Medicine (Bethesda, Md.) 2000; If the putative chromosomal location of a polynucleotide of the invention (Query sequence) was associated with a disease in the Morbid Map database, an OMIM reference identification number was noted in column 9, Table 1B.1, labeled “OMIM Disease Reference(s). Table 5 is a key to the OMIM reference identification numbers (column 1), and provides a description of the associated disease in Column 2

Table 1B.2

Column 5, in Table 1B.2, provides an expression profile and library code:count for each of the contig sequences (SEQ ID NO:X) disclosed in Table 1B, which can routinely be combined with the information provided in Table 4 and used to determine the tissues, cells, and/or cell line libraries which predominantly express the polynucleotides of the invention. The first number in Table 1B.2, column 5 (preceding the colon), represents the tissue/cell source identifier code corresponding to the code and description provided in Table 4. The second number in column 5 (following the colon) represents the number of times a sequence corresponding to the reference polynucleotide sequence was identified in the corresponding tissue/cell source. Those tissue/cell source identifier codes in which the first two letters are “AR” designate information generated using DNA array technology. Utilizing this technology, cDNAs were amplified by PCR and then transferred, in duplicate, onto the array. Gene expression was assayed through hybridization of first strand cDNA probes to the DNA array. cDNA probes were generated from total RNA extracted from a variety of different tissues and cell lines. Probe synthesis was performed in the presence of ³³P dCTP, using oligo (dT) to prime reverse transcription. After hybridization, high stringency washing conditions were employed to remove non-specific hybrids from the array. The remaining signal, emanating from each gene target, was measured using a Phosphorimager. Gene expression was reported as Phosphor Stimulating Luminescence (PSL) which reflects the level of phosphor signal generated from the probe hybridized to each of the gene targets represented on the array. A local background signal subtraction was performed before the total signal generated from each array was used to normalize gene expression between the different hybridizations. The value presented after “[array code]:” represents the mean of the duplicate values, following background subtraction and probe normalization. One of skill in the art could routinely use this information to identify normal and/or diseased tissue(s) which show a predominant expression pattern of the corresponding polynucleotide of the invention or to identify polynucleotides which show predominant and/or specific tissue and/or cell expression.

Lengthy table referenced here US20110105726A1-20110505-T00001 Please refer to the end of the specification for access instructions.

Lengthy table referenced here US20110105726A1-20110505-T00002 Please refer to the end of the specification for access instructions.

Table 1C summarizes additional polynucleotides encompassed by the invention (including cDNA clones related to the sequences (Clone ID:), contig sequences (contig identifier (Contig ID:) contig nucleotide sequence identifiers (SEQ ID NO:X)), and genomic sequences (SEQ ID NO:B). The first column provides a unique clone identifier, “Clone ID:”, for a cDNA clone related to each contig sequence. The second column provides the sequence identifier, “SEQ ID NO:X”, for each contig sequence. The third column provides a unique contig identifier, “Contig ID:” for each contig sequence. The fourth column, provides a BAC identifier “BAC ID NO:A” for the BAC clone referenced in the corresponding row of the table. The fifth column provides the nucleotide sequence identifier, “SEQ ID NO:B” for a fragment of the BAC clone identified in column four of the corresponding row of the table. The sixth column, “Exon From-To”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence of SEQ ID NO:B which delineate certain polynucleotides of the invention that are also exemplary members of polynucleotide sequences that encode polypeptides of the invention (e.g., polypeptides containing amino acid sequences encoded by the polynucleotide sequences delineated in column six, and fragments and variants thereof). Table 1C from U.S. patent application Ser. No. 10/472,532, filed Sep. 20, 2003, is herein incorporated by reference.

Tables 1D.1 and 1D.2: The polynucleotides or polypeptides, or agonists or antagonists of the present invention can be used in assays to test for one or more biological activities. If these polynucleotides and polypeptides do exhibit activity in a particular assay, it is likely that these molecules may be involved in the diseases associated with the biological activity. Thus, the polynucleotides or polypeptides, or agonists or antagonists could be used to treat the associated disease.

The present invention encompasses methods of detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating a disease or disorder. In preferred embodiments, the present invention encompasses a method of treating an immune disease or disorder comprising administering to a patient in which such detection, treatment, prevention, and/or amelioration is desired a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof) in an amount effective to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate the immune disease or disorder.

In another embodiment, the present invention also encompasses methods of detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating an immune disease or disorder; comprising administering to a patient combinations of the proteins, nucleic acids, or antibodies of the invention (or fragments or variants thereof), sharing similar indications as shown in the corresponding rows in Column 3 of Table 1E.

Tables 1D.1 and 1D.2 provide information related to biological activities for polynucleotides and polypeptides of the invention (including antibodies, agonists, and/or antagonists thereof). Tables 1D.1 and 1D.2 also provide information related to assays which may be used to test polynucleotides and polypeptides of the invention (including antibodies, agonists, and/or antagonists thereof) for the corresponding biological activities. The first column of Table 1D.1 (“Gene No.”) provides the gene number in the application for each clone identifier. The second column of Table 1D.1 (“cDNA Clone ID:”) provides the unique clone identifier for each clone as previously described and indicated in Table 1A through Table 1D. The third column of Table 1D.1 (“AA SEQ ID NO:Y”) indicates the Sequence Listing SEQ ID Number for polypeptide sequences encoded by the corresponding cDNA clones (also as indicated in Tables 1A, Table 1B, and Table 2). The fourth column of Table 1D.1 (“Biological Activity”) indicates a biological activity corresponding to the indicated polypeptides (or polynucleotides encoding said polypeptides). The first column of Table 1D.2 (“Biological Activity No.”) provides the biological activity number corresponding to each biological activity recited in the fourth column of Table 1D.1. The second column of Table 1D.2 (“Biological Activity”) indicates the biological activity corresponding to each biological activity number (also as indicated in Table 1D.1). The third column (“Exemplary Activity Assay”) further describes the corresponding biological activity and also provides information pertaining to the various types of assays which may be performed to test, demonstrate, or quantify the corresponding biological activity.

Tables 1D.1 and 1D.2 describe the use of, inter alia, FMAT technology for testing or demonstrating various biological activities. Fluorometric microvolume assay technology (FMAT) is a fluorescence-based system which provides a means to perform nonradioactive cell- and bead-based assays to detect activation of cell signal transduction pathways. This technology was designed specifically for ligand binding and immunological assays. Using this technology, fluorescent cells or beads at the bottom of the well are detected as localized areas of concentrated fluorescence using a data processing system. Unbound fluorophore comprising the background signal is ignored, allowing for a wide variety of homogeneous assays. FMAT technology may be used for peptide ligand binding assays, immunofluorescence, apoptosis, cytotoxicity, and bead-based immunocapture assays. See, Miraglia S et. al., “Homogeneous cell and bead based assays for high throughput screening using fluorometric microvolume assay technology,” Journal of Biomolecular Screening; 4:193-204 (1999). In particular, FMAT technology may be used to test, confirm, and/or identify the ability of polypeptides (including polypeptide fragments and variants) to activate signal transduction pathways. For example, FMAT technology may be used to test, confirm, and/or identify the ability of polypeptides to upregulate production of immunomodulatory proteins (such as, for example, interleukins, GM-CSF, Rantes, and Tumor Necrosis factors, as well as other cellular regulators (e.g. insulin)).

Tables 1D.1 and 1D.2 also describe the use of kinase assays for testing, demonstrating, or quantifying biological activity. In this regard, the phosphorylation and de-phosphorylation of specific amino acid residues (e.g. Tyrosine, Serine, Threonine) on cell-signal transduction proteins provides a fast, reversible means for activation and de-activation of cellular signal transduction pathways. Moreover, cell signal transduction via phosphorylation/de-phosphorylation is crucial to the regulation of a wide variety of cellular processes (e.g. proliferation, differentiation, migration, apoptosis, etc.). Accordingly, kinase assays provide a powerful tool useful for testing, confirming, and/or identifying polypeptides (including polypeptide fragments and variants) that mediate cell signal transduction events via protein phosphorylation. See e.g., Forrer, P., Tamaskovic R., and Jaussi, R. “Enzyme-Linked Immunosorbent Assay for Measurement of JNK, ERK, and p38 Kinase Activities” Biol. Chem. 379(8-9): 1101-1110 (1998). Table 1D from U.S. patent application Ser. No. 10/472,532, filed Sep. 20, 2003, is herein incorporated by reference.

TABLE 1D.1 AA Gene cDNA SEQ ID No. Clone ID NO: Y Biological Activity 1 H2CBG48 908 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 1 H2CBG48 908 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 1 H2CBG48 908 Biological Activity #21: Activation of transcription through cAMP response element in immune cells (such as T-cells). 2 H2MAC30 909 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 2 H2MAC30 909 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 3 H6EAB28 910 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 3 H6EAB28 910 Biological Activity #79: Production of TNF alpha by dendritic cells 4 H6EDF66 911 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 4 H6EDF66 911 Biological Activity #65: Production of IL-10 and activation of T-cells. 5 H6EDX46 912 Biological Activity #65: Production of IL-10 and activation of T-cells. 6 HABAG37 913 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 6 HABAG37 913 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 7 HACBD91 914 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 7 HACBD91 914 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #22: Activation of transcription through CD28 response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 7 HACBD91 914 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 7 HACBD91 914 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 7 HACBD91 914 Biological Activity #72: Production of IL-6 7 HACBD91 914 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 8 HACCI17 915 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 8 HACCI17 915 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 8 HACCI17 915 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 8 HACCI17 915 Biological Activity #60: Production of ICAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 8 HACCI17 915 Biological Activity #71: Production of IL-5 8 HACCI17 915 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 8 HACCI17 915 Biological Activity #75: Production of IL-8 by immune cells (such as the human EOL-1 eosinophil cells) 8 HACCI17 915 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 9 HADAO89 916 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 10 HADCP14 917 Biological Activity #78: Production of RANTES 11 HAGAI85 918 Biological Activity #59: Production of GM-CSF 11 HLTFA64 1210 Biological Activity #64: Production of IFNgamma using Natural Killer cells 12 HAGAM64 919 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 13 HAGAN21 920 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 14 HAGBZ81 921 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 16 HAGDS20 923 Biological Activity #76: Production of MCP-1 17 HAGFG51 924 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 18 HAHDB16 925 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 18 HAHDB16 925 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 18 HAHDB16 925 Biological Activity #63: Production of IFNgamma using a T cells 19 HAHDR32 926 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 20 HAIBO71 927 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 20 HAIBO71 927 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 20 HAIBO71 927 Biological Activity #56: Endothelial Cell Apoptosis 21 HAIBP89 928 Biological Activity #59: Production of GM-CSF 21 HAIBP89 928 Biological Activity #63: Production of IFNgamma using a T cells 22 HAICP19 929 Biological Activity #5: Activation of Adipocyte PI3 Kinase Signaling Pathway 22 HAICP19 929 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 22 HAICP19 929 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 22 HAICP19 929 Biological Activity #53: Bone marrow cell proliferation (fibronectin enhanced) 22 HAICP19 929 Biological Activity #61: Production of ICAM-1 23 HAIFL18 930 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 23 HAIFL18 930 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 23 HAIFL18 930 Biological Activity #63: Production of IFNgamma using a T cells 24 HAJAF57 931 Biological Activity #7: Activation of Endothelial Cell JNK Signaling Pathway. 24 HAJAF57 931 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 25 HAJBR69 932 Biological Activity #59: Production of GM-CSF 25 HAJBR69 932 Biological Activity #91: Regulation of transcription through the PEPCK promoter in hepatocytes 26 HAJBZ75 933 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 26 HAJBZ75 933 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 27 HAMFC93 934 Biological Activity #68: Production of IL-13 and activation of T-cells. 28 HAMFK58 935 Biological Activity #62: Production of ICAM-1 29 HAPNY86 936 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 30 HAPPW30 937 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 31 HAPQT22 938 Biological Activity #72: Production of IL-6 31 HAPQT22 938 Biological Activity #76: Production of MCP-1 32 HASAV70 939 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 32 HASAV70 939 Biological Activity #77: Production of MIP1alpha 33 HASCG84 940 Biological Activity #72: Production of IL-6 33 HASCG84 940 Biological Activity #77: Production of MIP1alpha 34 HATAC53 941 Biological Activity #59: Production of GM-CSF 35 HATBR65 942 Biological Activity #72: Production of IL-6 35 HATBR65 942 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 36 HATCB92 943 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 37 HATCP77 944 Biological Activity #72: Production of IL-6 37 HATCP77 944 Biological Activity #101: Upregulation of CD71 and activation of T cells 38 HATDF29 945 Biological Activity #65: Production of IL-10 and activation of T-cells. 39 HATDM46 946 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 39 HATDM46 946 Biological Activity #100: Upregulation of CD69 and activation of T cells 40 HATEE46 947 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 40 HATEE46 947 Biological Activity #62: Production of ICAM-1 41 HBAFJ33 948 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 41 HBAFJ33 948 Biological Activity #98: Upregulation of CD152 and activation of T cells 42 HBAFV19 949 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 42 HBAFV19 949 Biological Activity #98: Upregulation of CD152 and activation of T cells 43 HBAMB34 950 Biological Activity #100: Upregulation of CD69 and activation of T cells 43 HBAMB34 950 Biological Activity #101: Upregulation of CD71 and activation of T cells 44 HBCPB32 951 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 45 HBHAD12 952 Biological Activity #63: Production of IFNgamma using a T cells 46 HBHMA23 953 Biological Activity #79: Production of TNF alpha by dendritic cells 47 HBIBW67 954 Biological Activity #71: Production of IL-5 48 HBIMB51 955 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 49 HBINS58 956 Biological Activity #57: Insulin Secretion 49 HBINS58 956 Biological Activity #79: Production of TNF alpha by dendritic cells 50 HBJFU48 957 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 50 HBJFU48 957 Biological Activity #72: Production of IL-6 51 HBJID05 958 Biological Activity #65: Production of IL-10 and activation of T-cells. 52 HBJIY92 959 Biological Activity #72: Production of IL-6 52 HBJIY92 959 Biological Activity #79: Production of TNF alpha by dendritic cells 53 HBJJU28 960 Biological Activity #68: Production of IL-13 and activation of T-cells. 54 HBJLC01 961 Biological Activity #5: Activation of Adipocyte PI3 Kinase Signaling Pathway 54 HBJLC01 961 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 54 HBJLC01 961 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 54 HBJLC01 961 Biological Activity #50: Activation of transcription through STAT6 response element in immune cells (such as mast cells). 54 HBJLC01 961 Biological Activity #72: Production of IL-6 55 HBJLF01 962 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 55 HBJLF01 962 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 56 HBJLH40 963 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 57 HBJNC59 964 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 58 HBNAW17 965 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 58 HBNAW17 965 Biological Activity #57: Insulin Secretion 59 HBOEG11 966 Biological Activity #98: Upregulation of CD152 and activation of T cells 60 HBOEG69 967 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 61 HBXFL29 968 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 62 HCACU58 969 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 62 HCACU58 969 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 62 HCACU58 969 Biological Activity #60: Production of ICAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 62 HCACU58 969 Biological Activity #65: Production of IL-10 and activation of T-cells. 63 HCACV51 970 Biological Activity #69: Production of IL-2 and activation of T cells 64 HCDBW86 971 Biological Activity #71: Production of IL-5 65 HCE1Q89 972 Biological Activity #63: Production of IFNgamma using a T cells 66 HCE2F54 973 Biological Activity #36: Activation of transcription through NFKB response element in epithelial cells (such as HELA cells). 66 HCE2F54 973 Biological Activity #44: Activation of transcription through NFKB response element in immune cells (such as the U937 human monocyte cell line). 66 HCE2F54 973 Biological Activity #91: Regulation of transcription through the PEPCK promoter in hepatocytes 67 HCE3G69 974 Biological Activity #65: Production of IL-10 and activation of T-cells. 67 HCE3G69 974 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 68 HCEEA88 975 Biological Activity #65: Production of IL-10 and activation of T-cells. 69 HCEFB69 976 Biological Activity #25: Activation of transcription through GAS response element in immune cells (such as eosinophils). 69 HCEFB69 976 Biological Activity #65: Production of IL-10 and activation of T-cells. 70 HCEFB80 977 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 70 HCEFB80 977 Biological Activity #57: Insulin Secretion 71 HCEGR33 978 Biological Activity #62: Production of ICAM-1 72 HCEMP62 979 Biological Activity #36: Activation of transcription through NFKB response element in epithelial cells (such as HELA cells). 72 HCEMP62 979 Biological Activity #44: Activation of transcription through NFKB response element in immune cells (such as the U937 human monocyte cell line). 73 HCENK38 980 Biological Activity #9: Activation of Hepatocyte ERK Signaling Pathway 73 HCENK38 980 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 73 HCENK38 980 Biological Activity #83: Protection from Endothelial Cell Apoptosis. 73 HCENK38 980 Biological Activity #101: Upregulation of CD71 and activation of T cells 74 HCEWE17 981 Biological Activity #62: Production of ICAM-1 75 HCEWE20 982 Biological Activity #61: Production of ICAM-1 75 HCEWE20 982 Biological Activity #89: Regulation of transcription of Malic Enzyme in hepatocytes 76 HCFCU88 983 Biological Activity #98: Upregulation of CD152 and activation of T cells 77 HCFMV71 984 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 77 HCFMV71 984 Biological Activity #20: Activation of transcription through cAMP response element in immune cells (such as T-cells). 77 HCFMV71 984 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 77 HCFMV71 984 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 78 HCFNN01 985 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 79 HCFOM18 986 Biological Activity #98: Upregulation of CD152 and activation of T cells 79 HCFOM18 986 Biological Activity #100: Upregulation of CD69 and activation of T cells 79 HCFOM18 986 Biological Activity #101: Upregulation of CD71 and activation of T cells 79 HCFOM18 986 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 80 HCHNF25 987 Biological Activity #55: Calcium flux in immune cells (such as monocytes) 81 HCMSQ56 988 Biological Activity #62: Production of ICAM-1 82 HCMST14 989 Biological Activity #72: Production of IL-6 83 HCMTB45 990 Biological Activity #98: Upregulation of CD152 and activation of T cells 84 HCNSD93 991 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 85 HCOOS80 992 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 86 HCQCT05 993 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 87 HCUBS50 994 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 87 HCUBS50 994 Biological Activity #25: Activation of transcription through GAS response element in immune cells (such as eosinophils). 88 HCUCK44 995 Biological Activity #76: Production of MCP-1 88 HCUCK44 995 Biological Activity #83: Protection from Endothelial Cell Apoptosis. 89 HCUEO60 996 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 90 HCUGM86 997 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 91 HCUHK65 998 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 92 HCUIM65 999 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 92 HCUIM65 999 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 92 HCUIM65 999 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 92 HCUIM65 999 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 92 HCUIM65 999 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 92 HCUIM65 999 Biological Activity #39: Activation of transcription through NFKB response element in immune cells (such as mast cells). 92 HCUIM65 999 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 92 HCUIM65 999 Biological Activity #46: Activation of transcription through serum response element in immune cells (such as T-cells). 92 HCUIM65 999 Biological Activity #49: Activation of transcription through serum response element in pre-adipocytes. 92 HCUIM65 999 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 92 HCUIM65 999 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 92 HCUIM65 999 Biological Activity #96: Stimulation of Calcium Flux in pancreatic beta cells. 93 HCWEB58 1000 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 94 HCWGU37 1001 Biological Activity #54: Calcium flux in chondrocytes 95 HCWKC15 1002 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 95 HCWKC15 1002 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 95 HCWKC15 1002 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 95 HCWKC15 1002 Biological Activity #25: Activation of transcription through GAS response element in immune cells (such as eosinophils). 95 HCWKC15 1002 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 95 HCWKC15 1002 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 95 HCWKC15 1002 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 95 HCWKC15 1002 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 95 HCWKC15 1002 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 95 HCWKC15 1002 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 95 HCWKC15 1002 Biological Activity #38: Activation of transcription through NFKB response element in immune cells (such as EOL1 cells). 95 HCWKC15 1002 Biological Activity #39: Activation of transcription through NFKB response element in immune cells (such as mast cells). 95 HCWKC15 1002 Biological Activity #40: Activation of transcription through NFKB response element in immune cells (such as natural killer cells). 95 HCWKC15 1002 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 95 HCWKC15 1002 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 95 HCWKC15 1002 Biological Activity #46: Activation of transcription through serum response element in immune cells (such as T-cells). 95 HCWKC15 1002 Biological Activity #49: Activation of transcription through serum response element in pre-adipocytes. 95 HCWKC15 1002 Biological Activity #50: Activation of transcription through STAT6 response element in immune cells (such as mast cells). 95 HCWKC15 1002 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 95 HCWKC15 1002 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 96 HCWLD74 1003 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 96 HCWLD74 1003 Biological Activity #21: Activation of transcription through cAMP response element in immune cells (such as T-cells). 96 HCWLD74 1003 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 96 HCWLD74 1003 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 96 HCWLD74 1003 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 96 HCWLD74 1003 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 96 HCWLD74 1003 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 96 HCWLD74 1003 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 97 HDHEB60 1004 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 97 HDHEB60 1004 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 97 HDHEB60 1004 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 97 HDHEB60 1004 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 97 HDHEB60 1004 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 97 HDHEB60 1004 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 97 HDHEB60 1004 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 97 HDHEB60 1004 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 97 HDHEB60 1004 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 97 HDHEB60 1004 Biological Activity #58: Myoblast cell proliferation 97 HDHEB60 1004 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 98 HDHIA94 1005 Biological Activity #79: Production of TNF alpha by dendritic cells 99 HDHMA45 1006 Biological Activity #65: Production of IL-10 and activation of T-cells. 100 HDHMA72 1007 Biological Activity #40: Activation of transcription through NFKB response element in immune cells (such as natural killer cells). 101 HDLAC10 1008 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 101 HDLAC10 1008 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 102 HDLAO28 1009 Biological Activity #98: Upregulation of CD152 and activation of T cells 103 HDPBA28 1010 Biological Activity #65: Production of IL-10 and activation of T-cells. 103 HDPBA28 1010 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 104 HDPBQ02 1011 Biological Activity #100: Upregulation of CD69 and activation of T cells 105 HDPBQ71 1012 Biological Activity #63: Production of IFNgamma using a T cells 105 HDPBQ71 1012 Biological Activity #95: Regulation of viability or proliferation of immune cells (such as human eosinophil EOL-1 cells). 106 HDPCO25 1013 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 106 HDPCO25 1013 Biological Activity #94: Regulation of viability and proliferation of pancreatic beta cells. 107 HDPCY37 1014 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 107 HDPCY37 1014 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 107 HDPCY37 1014 Biological Activity #65: Production of IL-10 and activation of T-cells. 108 HDPFF39 1015 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 109 HDPGK25 1016 Biological Activity #76: Production of MCP-1 110 HDPGP94 1017 Biological Activity #77: Production of MIP1alpha 110 HDPGP94 1017 Biological Activity #79: Production of TNF alpha by dendritic cells 111 HDPHI51 1018 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 111 HDPHI51 1018 Biological Activity #90: Regulation of transcription through the FAS promoter element in hepatocytes 112 HDPJF37 1019 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 113 HDPJM30 1020 Biological Activity #76: Production of MCP-1 113 HDPJM30 1020 Biological Activity #90: Regulation of transcription through the FAS promoter element in hepatocytes 114 HDPNC61 1021 Biological Activity #6: Activation of Endothelial Cell ERK Signaling Pathway. 114 HDPNC61 1021 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 114 HDPNC61 1021 Biological Activity #25: Activation of transcription through GAS response element in immune cells (such as eosinophils). 114 HDPNC61 1021 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 115 HDPND46 1022 Biological Activity #5: Activation of Adipocyte PI3 Kinase Signaling Pathway 115 HDPND46 1022 Biological Activity #70: Production of IL-4 115 HDPND46 1022 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 116 HDPOE32 1023 Biological Activity #62: Production of ICAM-1 117 HDPOH06 1024 Biological Activity #62: Production of ICAM-1 117 HDPOH06 1024 Biological Activity #65: Production of IL-10 and activation of T-cells. 118 HDPOZ56 1025 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 118 HDPOZ56 1025 Biological Activity #24: Activation of transcription through GAS response element in epithelial cells (such as HELA cells). 119 HDPPA04 1026 Biological Activity #77: Production of MIP1alpha 120 HDPPH47 1027 Biological Activity #76: Production of MCP-1 121 HDPSB18 1028 Biological Activity #66: Production of IL-10 and downregulation of immune responses 121 HDPSB18 1028 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 122 HDPSP01 1029 Biological Activity #57: Insulin Secretion 122 HDPSP01 1029 Biological Activity #76: Production of MCP-1 123 HDPSP54 1030 Biological Activity #7: Activation of Endothelial Cell JNK Signaling Pathway. 123 HDPSP54 1030 Biological Activity #65: Production of IL-10 and activation of T-cells. 123 HDPSP54 1030 Biological Activity #85: Regulation of apoptosis in pancreatic beta cells. 124 HDPSU13 1031 Biological Activity #76: Production of MCP-1 125 HDPTD15 1032 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 125 HDPTD15 1032 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 125 HDPTD15 1032 Biological Activity #22: Activation of transcription through CD28 response element in immune cells (such as T-cells). 125 HDPTD15 1032 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 125 HDPTD15 1032 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 126 HDPTK41 1033 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 126 HDPTK41 1033 Biological Activity #63: Production of IFNgamma using a T cells 127 HDPUG50 1034 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 128 HDPUH26 1035 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 128 HDPUH26 1035 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 129 HDPUW68 1036 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 129 HDPUW68 1036 Biological Activity #12: Activation of Skeletal Muscle Cell PI3 Kinase Signaling Pathway 129 HDPUW68 1036 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 129 HDPUW68 1036 Biological Activity #96: Stimulation of Calcium Flux in pancreatic beta cells. 130 HDPVH60 1037 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 131 HDPWN93 1038 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 131 HDPWN93 1038 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 131 HDPWN93 1038 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 132 HDQHD03 1039 Biological Activity #65: Production of IL-10 and activation of T-cells. 132 HDQHD03 1039 Biological Activity #98: Upregulation of CD152 and activation of T cells 133 HDTBP04 1040 Biological Activity #62: Production of ICAM-1 133 HDTBP04 1040 Biological Activity #72: Production of IL-6 133 HDTBP04 1040 Biological Activity #76: Production of MCP-1 134 HAGAI85 918 Biological Activity #64: Production of IFNgamma using Natural Killer cells 135 HDTEN81 1042 Biological Activity #98: Upregulation of CD152 and activation of T cells 136 HDTFE17 1043 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 137 HDTGC73 1044 Biological Activity #62: Production of ICAM-1 138 HDTIT10 1045 Biological Activity #72: Production of IL-6 139 HDTMK50 1046 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 140 HE2DY70 1047 Biological Activity #63: Production of IFNgamma using a T cells 141 HE2EN04 1048 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 142 HE2FV03 1049 Biological Activity #61: Production of ICAM-1 143 HE2NV57 1050 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 143 HE2NV57 1050 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 143 HE2NV57 1050 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 143 HE2NV57 1050 Biological Activity #22: Activation of transcription through CD28 response element in immune cells (such as T-cells). 143 HE2NV57 1050 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 143 HE2NV57 1050 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 143 HE2NV57 1050 Biological Activity #57: Insulin Secretion 144 HE2PD49 1051 Biological Activity #72: Production of IL-6 145 HE2PY40 1052 Biological Activity #98: Upregulation of CD152 and activation of T cells 146 HE6EU50 1053 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 146 HE6EU50 1053 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 146 HE6EU50 1053 Biological Activity #100: Upregulation of CD69 and activation of T cells 147 HE8MH91 1054 Biological Activity #2: Activation of transcription through NFKB response element in immune cells (such as B-cells). 148 HE8QV67 1055 Biological Activity #70: Production of IL-4 149 HE8UB86 1056 Biological Activity #65: Production of IL-10 and activation of T-cells. 150 HE9BK23 1057 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 150 HE9BK23 1057 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 151 HE9CO69 1058 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 152 HE9CP41 1059 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 153 HE9DG49 1060 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 153 HE9DG49 1060 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 153 HE9DG49 1060 Biological Activity #22: Activation of transcription through CD28 response element in immune cells (such as T-cells). 153 HE9DG49 1060 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 153 HE9DG49 1060 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 154 HE9OW20 1061 Biological Activity #13: Activation of Skeletal Muscle Cell ERK Signaling Pathway 154 HE9OW20 1061 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 155 HE9RM63 1062 Biological Activity #36: Activation of transcription through NFKB response element in epithelial cells (such as HELA cells). 156 HEAAR07 1063 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 157 HEBAE88 1064 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 158 HEBBN36 1065 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 159 HEBCM63 1066 Biological Activity #25: Activation of transcription through GAS response element in immune cells (such as eosinophils). 159 HEBCM63 1066 Biological Activity #63: Production of IFNgamma using a T cells 160 HEBEJ18 1067 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 161 HEEAG23 1068 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 161 HEEAG23 1068 Biological Activity #12: Activation of Skeletal Muscle Cell PI3 Kinase Signaling Pathway 161 HEEAG23 1068 Biological Activity #100: Upregulation of CD69 and activation of T cells 162 HEEAJ02 1069 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 163 HEEAQ11 1070 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 163 HEEAQ11 1070 Biological Activity #95: Regulation of viability or proliferation of immune cells (such as human eosinophil EOL-1 cells). 163 HEEAQ11 1070 Biological Activity #101: Upregulation of CD71 and activation of T cells 164 HEGAN94 1071 Biological Activity #62: Production of ICAM-1 165 HEGBS69 1072 Biological Activity #72: Production of IL-6 165 HEGBS69 1072 Biological Activity #98: Upregulation of CD152 and activation of T cells 166 HELGK31 1073 Biological Activity #63: Production of IFNgamma using a T cells 166 HELGK31 1073 Biological Activity #72: Production of IL-6 167 HELHD85 1074 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 168 HELHL48 1075 Biological Activity #98: Upregulation of CD152 and activation of T cells 169 HEMAM41 1076 Biological Activity #72: Production of IL-6 169 HEMAM41 1076 Biological Activity #79: Production of TNF alpha by dendritic cells 170 HEPAA46 1077 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 171 HEQAK71 1078 Biological Activity #62: Production of ICAM-1 171 HEQAK71 1078 Biological Activity #79: Production of TNF alpha by dendritic cells 172 HEQCC55 1079 Biological Activity #68: Production of IL-13 and activation of T-cells. 172 HEQCC55 1079 Biological Activity #76: Production of MCP-1 173 HERAD40 1080 Biological Activity #100: Upregulation of CD69 and activation of T cells 174 HERAR44 1081 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 174 HERAR44 1081 Biological Activity #62: Production of ICAM-1 175 HESAJ10 1082 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 176 HETAB45 1083 Biological Activity #2: Activation of transcription through NFKB response element in immune cells (such as B-cells). 177 HETBR16 1084 Biological Activity #62: Production of ICAM-1 178 HETEU28 1085 Biological Activity #71: Production of IL-5 179 HETLM70 1086 Biological Activity #72: Production of IL-6 179 HETLM70 1086 Biological Activity #77: Production of MIP1alpha 179 HETLM70 1086 Biological Activity #79: Production of TNF alpha by dendritic cells 180 HFABG18 1087 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 180 HFABG18 1087 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 180 HFABG18 1087 Biological Activity #63: Production of IFNgamma using a T cells 180 HFABG18 1087 Biological Activity #83: Protection from Endothelial Cell Apoptosis. 181 HFABH95 1088 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 181 HFABH95 1088 Biological Activity #100: Upregulation of CD69 and activation of T cells 182 HFAMB72 1089 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 183 HFAMH77 1090 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 183 HFAMH77 1090 Biological Activity #63: Production of IFNgamma using a T cells 184 HFCCQ50 1091 Biological Activity #26: Activation of transcription through GAS response element in immune cells (such as monocytes). 184 HFCCQ50 1091 Biological Activity #43: Activation of transcription through NFKB response element in immune cells (such as the Jurkat human T cell line). 184 HFCCQ50 1091 Biological Activity #70: Production of IL-4 184 HFCCQ50 1091 Biological Activity #79: Production of TNF alpha by dendritic cells 185 HFCDK17 1092 Biological Activity #76: Production of MCP-1 187 HFFAD59 1094 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 187 HFFAD59 1094 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 187 HFFAD59 1094 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 188 HFFAL36 1095 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 188 HFFAL36 1095 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 189 HFGAD82 1096 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 189 HFGAD82 1096 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 190 HFIIN69 1097 Biological Activity #66: Production of IL-10 and downregulation of immune responses 190 HFIIN69 1097 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 191 HFIIZ70 1098 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 192 HFKET18 1099 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 192 HFKET18 1099 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 192 HFKET18 1099 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 193 HFLNB64 1100 Biological Activity #71: Production of IL-5 194 HFOXA73 1101 Biological Activity #65: Production of IL-10 and activation of T-cells. 195 HFOXB13 1102 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 196 HFPAC12 1103 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 197 HFPAO71 1104 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 197 HFPAO71 1104 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 197 HFPAO71 1104 Biological Activity #75: Production of IL-8 by immune cells (such as the human EOL-1 eosinophil cells) 198 HFPCX09 1105 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 198 HFPCX09 1105 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 198 HFPCX09 1105 Biological Activity #79: Production of TNF alpha by dendritic cells 199 HFPCX36 1106 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 200 HFPCX64 1107 Biological Activity #71: Production of IL-5 201 HFRAN90 1108 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 201 HFRAN90 1108 Biological Activity #62: Production of ICAM-1 202 HFTBM50 1109 Biological Activity #57: Insulin Secretion 202 HFTBM50 1109 Biological Activity #65: Production of IL-10 and activation of T-cells. 203 HFTDL56 1110 Biological Activity #62: Production of ICAM-1 205 HFXAM76 1112 Biological Activity #59: Production of GM-CSF 206 HFXDJ75 1113 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 206 HFXDJ75 1113 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 206 HFXDJ75 1113 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 207 HFXDN63 1114 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 208 HFXGT26 1115 Biological Activity #62: Production of ICAM-1 209 HFXGV31 1116 Biological Activity #79: Production of TNF alpha by dendritic cells 210 HFXHD88 1117 Biological Activity #98: Upregulation of CD152 and activation of T cells 211 HFXJU68 1118 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 211 HFXJU68 1118 Biological Activity #65: Production of IL-10 and activation of T-cells. 212 HFXKJ03 1119 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 213 HFXKY27 1120 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 213 HFXKY27 1120 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 214 HGBFO79 1121 Biological Activity #61: Production of ICAM-1 214 HGBFO79 1121 Biological Activity #82: Proliferation of immune cells (such as the HMC-1 human mast cell line) 215 HGBHE57 1122 Biological Activity #101: Upregulation of CD71 and activation of T cells 216 HGBIB74 1123 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 216 HGBIB74 1123 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 216 HGBIB74 1123 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 216 HGBIB74 1123 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 217 HGLAL82 1124 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 218 HHAAF20 1125 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 219 HHEAA08 1126 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 219 HHEAA08 1126 Biological Activity #78: Production of RANTES 219 HHEAA08 1126 Biological Activity #98: Upregulation of CD152 and activation of T cells 220 HHEBB10 1127 Biological Activity #77: Production of MIP1alpha 221 HHEMA59 1128 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 222 HHEMA75 1129 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 222 HHEMA75 1129 Biological Activity #20: Activation of transcription through cAMP response element in immune cells (such as T-cells). 222 HHEMA75 1129 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 222 HHEMA75 1129 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 222 HHEMA75 1129 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 222 HHEMA75 1129 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 222 HHEMA75 1129 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 222 HHEMA75 1129 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 223 HHEMM74 1130 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 224 HHENK42 1131 Biological Activity #68: Production of IL-13 and activation of T-cells. 225 HHENP27 1132 Biological Activity #80: Production of TNF alpha by T cells 226 HHENQ22 1133 Biological Activity #72: Production of IL-6 227 HHEPD24 1134 Biological Activity #72: Production of IL-6 227 HHEPD24 1134 Biological Activity #76: Production of MCP-1 227 HHEPD24 1134 Biological Activity #77: Production of MIP1alpha 227 HHEPD24 1134 Biological Activity #79: Production of TNF alpha by dendritic cells 228 HHEPM33 1135 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 228 HHEPM33 1135 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 228 HHEPM33 1135 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 228 HHEPM33 1135 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 228 HHEPM33 1135 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 228 HHEPM33 1135 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 228 HHEPM33 1135 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 229 HHEPT60 1136 Biological Activity #40: Activation of transcription through NFKB response element in immune cells (such as natural killer cells). 230 HHEPU04 1137 Biological Activity #72: Production of IL-6 230 HHEPU04 1137 Biological Activity #79: Production of TNF alpha by dendritic cells 231 HHFEC49 1138 Biological Activity #63: Production of IFNgamma using a T cells 232 HHFGR93 1139 Biological Activity #36: Activation of transcription through NFKB response element in epithelial cells (such as HELA cells). 232 HHFGR93 1139 Biological Activity #55: Calcium flux in immune cells (such as monocytes) 233 HHFHJ59 1140 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 233 HHFHJ59 1140 Biological Activity #65: Production of IL-10 and activation of T-cells. 233 HHFHJ59 1140 Biological Activity #100: Upregulation of CD69 and activation of T cells 233 HHFHJ59 1140 Biological Activity #101: Upregulation of CD71 and activation of T cells 233 HHFHJ59 1140 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 234 HHFHR32 1141 Biological Activity #72: Production of IL-6 235 HHFOJ29 1142 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 236 HHGCM76 1143 Biological Activity #62: Production of ICAM-1 236 HHGCM76 1143 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 237 HHGDF16 1144 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 237 HHGDF16 1144 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 238 HHGDW43 1145 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 238 HHGDW43 1145 Biological Activity #65: Production of IL-10 and activation of T-cells. 239 HHPEC09 1146 Biological Activity #65: Production of IL-10 and activation of T-cells. 240 HHPGO40 1147 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 240 HHPGO40 1147 Biological Activity #82: Proliferation of immune cells (such as the HMC-1 human mast cell line) 240 HHPGO40 1147 Biological Activity #100: Upregulation of CD69 and activation of T cells 241 HHPTJ65 1148 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 242 HHSDX28 1149 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 242 HHSDX28 1149 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 242 HHSDX28 1149 Biological Activity #79: Production of TNF alpha by dendritic cells 243 HHSGW69 1150 Biological Activity #66: Production of IL-10 and downregulation of immune responses 244 HHTLF25 1151 Biological Activity #66: Production of IL-10 and downregulation of immune responses 245 HJABX32 1152 Biological Activity #65: Production of IL-10 and activation of T-cells. 246 HJACA79 1153 Biological Activity #76: Production of MCP-1 248 HJACG30 1155 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 248 HJACG30 1155 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 249 HJBAV55 1156 Biological Activity #77: Production of MIP1alpha 250 HJBCU04 1157 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 250 HJBCU04 1157 Biological Activity #70: Production of IL-4 251 HJMBI18 1158 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 252 HJMBN89 1159 Biological Activity #65: Production of IL-10 and activation of T-cells. 253 HJMBT65 1160 Biological Activity #77: Production of MIP1alpha 253 HJMBT65 1160 Biological Activity #101: Upregulation of CD71 and activation of T cells 254 HJMBW30 1161 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 255 HJPAD75 1162 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 255 HJPAD75 1162 Biological Activity #72: Production of IL-6 255 HJPAD75 1162 Biological Activity #90: Regulation of transcription through the FAS promoter element in hepatocytes 256 HKAAE44 1163 Biological Activity #98: Upregulation of CD152 and activation of T cells 257 HKAAH36 1164 Biological Activity #72: Production of IL-6 257 HKAAH36 1164 Biological Activity #76: Production of MCP-1 258 HKAAK02 1165 Biological Activity #72: Production of IL-6 258 HKAAK02 1165 Biological Activity #76: Production of MCP-1 259 HKABI84 1166 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 259 HKABI84 1166 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 259 HKABI84 1166 Biological Activity #56: Endothelial Cell Apoptosis 260 HKABZ65 1167 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 260 HKABZ65 1167 Biological Activity #72: Production of IL-6 260 HKABZ65 1167 Biological Activity #85: Regulation of apoptosis in pancreatic beta cells. 261 HKACB56 1168 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 261 HKACB56 1168 Biological Activity #58: Myoblast cell proliferation 261 HKACB56 1168 Biological Activity #71: Production of IL-5 261 HKACB56 1168 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 261 HKACB56 1168 Biological Activity #98: Upregulation of CD152 and activation of T cells 262 HKACD58 1169 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 262 HKACD58 1169 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 263 HKACM93 1170 Biological Activity #25: Activation of transcription through GAS response element in immune cells (such as eosinophils). 263 HKACM93 1170 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 263 HKACM93 1170 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 264 HKADQ91 1171 Biological Activity #65: Production of IL-10 and activation of T-cells. 265 HKAEG43 1172 Biological Activity #71: Production of IL-5 266 HKAEL80 1173 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 267 HKAEV06 1174 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 267 HKAEV06 1174 Biological Activity #94: Regulation of viability and proliferation of pancreatic beta cells. 268 HKAFK41 1175 Biological Activity #61: Production of ICAM-1 268 HKAFK41 1175 Biological Activity #72: Production of IL-6 269 HKDBF34 1176 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 270 HKGAT94 1177 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 271 HKGCO27 1178 Biological Activity #59: Production of GM-CSF 272 HKISB57 1179 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 272 HKISB57 1179 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 273 HKIYH57 1180 Biological Activity #77: Production of MIP1alpha 274 HKIYP40 1181 Biological Activity #65: Production of IL-10 and activation of T-cells. 275 HKMLK53 1182 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 276 HKMLP68 1183 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 278 HL2AG57 1185 Biological Activity #72: Production of IL-6 279 HLCND09 1186 Biological Activity #98: Upregulation of CD152 and activation of T cells 280 HLDBX13 1187 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 280 HLDBX13 1187 Biological Activity #79: Production of TNF alpha by dendritic cells 281 HLDON23 1188 Biological Activity #62: Production of ICAM-1 281 HLDON23 1188 Biological Activity #65: Production of IL-10 and activation of T-cells. 281 HLDON23 1188 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 281 HLDON23 1188 Biological Activity #91: Regulation of transcription through the PEPCK promoter in hepatocytes 282 HLDOW79 1189 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 282 HLDOW79 1189 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 282 HLDOW79 1189 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 282 HLDOW79 1189 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 282 HLDOW79 1189 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 282 HLDOW79 1189 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 283 HLDQC46 1190 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 284 HLDQR62 1191 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 284 HLDQR62 1191 Biological Activity #93: Regulation of viability and proliferation of pancreatic beta cells. 285 HLDQU79 1192 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 285 HLDQU79 1192 Biological Activity #93: Regulation of viability and proliferation of pancreatic beta cells. 286 HLDRM43 1193 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 286 HLDRM43 1193 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 286 HLDRM43 1193 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 286 HLDRM43 1193 Biological Activity #98: Upregulation of CD152 and activation of T cells 287 HLDRP33 1194 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 288 HLHFP03 1195 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 289 HLHFR58 1196 Biological Activity #78: Production of RANTES 289 HLHFR58 1196 Biological Activity #79: Production of TNF alpha by dendritic cells 290 HLIBD68 1197 Biological Activity #72: Production of IL-6 290 HLIBD68 1197 Biological Activity #77: Production of MIP1alpha 290 HLIBD68 1197 Biological Activity #79: Production of TNF alpha by dendritic cells 290 HLIBD68 1197 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 291 HLICQ90 1198 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 291 HLICQ90 1198 Biological Activity #79: Production of TNF alpha by dendritic cells 291 HLICQ90 1198 Biological Activity #96: Stimulation of Calcium Flux in pancreatic beta cells. 291 HLICQ90 1198 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 292 HLJBJ61 1199 Biological Activity #31: Activation of transcription through GATA-3 response element in immune cells (such as T-cells). 293 HLMBO76 1200 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 294 HLMCA59 1201 Biological Activity #76: Production of MCP-1 295 HLQBE09 1202 Biological Activity #75: Production of IL-8 by immune cells (such as the human EOL-1 eosinophil cells) 295 HLQBE09 1202 Biological Activity #101: Upregulation of CD71 and activation of T cells 296 HLQDH79 1203 Biological Activity #65: Production of IL-10 and activation of T-cells. 297 HLQDR48 1204 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 297 HLQDR48 1204 Biological Activity #76: Production of MCP-1 297 HLQDR48 1204 Biological Activity #79: Production of TNF alpha by dendritic cells 298 HLQEM64 1205 Biological Activity #66: Production of IL-10 and downregulation of immune responses 299 HLTAU74 1206 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 299 HLTAU74 1206 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 300 HLTCO33 1207 Biological Activity #66: Production of IL-10 and downregulation of immune responses 301 HLTDV50 1208 Biological Activity #62: Production of ICAM-1 302 HLTEJ06 1209 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 303 HDTEK44 1041 Biological Activity #64: Production of IFNgamma using Natural Killer cells 304 HLTHG37 1211 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 305 HLWAA17 1212 Biological Activity #62: Production of ICAM-1 305 HLWAA17 1212 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 306 HLWAD77 1213 Biological Activity #1: Activation of transcription through the EGR (Early Growth Response) element in immune cells (such as B-cells). 307 HLWAE11 1214 Biological Activity #40: Activation of transcription through NFKB response element in immune cells (such as natural killer cells). 307 HLWAE11 1214 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 307 HLWAE11 1214 Biological Activity #55: Calcium flux in immune cells (such as monocytes) 308 HLWAO22 1215 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 308 HLWAO22 1215 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 308 HLWAO22 1215 Biological Activity #76: Production of MCP-1 309 HLWAY54 1216 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 309 HLWAY54 1216 Biological Activity #76: Production of MCP-1 309 HLWAY54 1216 Biological Activity #98: Upregulation of CD152 and activation of T cells 310 HLWBI63 1217 Biological Activity #101: Upregulation of CD71 and activation of T cells 311 HLWBY76 1218 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 311 HLWBY76 1218 Biological Activity #98: Upregulation of CD152 and activation of T cells 311 HLWBY76 1218 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 312 HLWCF05 1219 Biological Activity #5: Activation of Adipocyte PI3 Kinase Signaling Pathway 312 HLWCF05 1219 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 312 HLWCF05 1219 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 312 HLWCF05 1219 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 312 HLWCF05 1219 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 312 HLWCF05 1219 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 313 HLYAC95 1220 Biological Activity #63: Production of IFNgamma using a T cells 313 HLYAC95 1220 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 314 HLYAF80 1221 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 315 HLYAN59 1222 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 315 HLYAN59 1222 Biological Activity #98: Upregulation of CD152 and activation of T cells 315 HLYAN59 1222 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 316 HLYAZ61 1223 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 317 HLYBD32 1224 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 318 HMADS41 1225 Biological Activity #9: Activation of Hepatocyte ERK Signaling Pathway 318 HMADS41 1225 Biological Activity #83: Protection from Endothelial Cell Apoptosis. 318 HMADS41 1225 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 319 HMADU73 1226 Biological Activity #65: Production of IL-10 and activation of T-cells. 319 HMADU73 1226 Biological Activity #72: Production of IL-6 319 HMADU73 1226 Biological Activity #79: Production of TNF alpha by dendritic cells 320 HMAMI15 1227 Biological Activity #96: Stimulation of Calcium Flux in pancreatic beta cells. 320 HMAMI15 1227 Biological Activity #98: Upregulation of CD152 and activation of T cells 321 HMDAE65 1228 Biological Activity #72: Production of IL-6 322 HMDAN54 1229 Biological Activity #78: Production of RANTES 323 HMDAQ29 1230 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 323 HMDAQ29 1230 Biological Activity #61: Production of ICAM-1 324 HMEAI48 1231 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 325 HMECK83 1232 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 326 HMEED18 1233 Biological Activity #73: Production of IL6 by primary human aortic smooth muscle or normal human dermal fibroblast cells (without or with costimulation with TNFalpha). 326 HMEED18 1233 Biological Activity #96: Stimulation of Calcium Flux in pancreatic beta cells. 326 HMEED18 1233 Biological Activity #100: Upregulation of CD69 and activation of T cells 327 HMEET96 1234 Biological Activity #60: Production of ICAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 327 HMEET96 1234 Biological Activity #72: Production of IL-6 328 HMIAL37 1235 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 328 HMIAL37 1235 Biological Activity #65: Production of IL-10 and activation of T-cells. 329 HMIAP86 1236 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 329 HMIAP86 1236 Biological Activity #75: Production of IL-8 by immune cells (such as the human EOL-1 eosinophil cells) 329 HMIAP86 1236 Biological Activity #76: Production of MCP-1 329 HMIAP86 1236 Biological Activity #77: Production of MIP1alpha 329 HMIAP86 1236 Biological Activity #79: Production of TNF alpha by dendritic cells 329 HMIAP86 1236 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 330 HMKCG09 1237 Biological Activity #63: Production of IFNgamma using a T cells 330 HMKCG09 1237 Biological Activity #65: Production of IL-10 and activation of T-cells. 330 HMKCG09 1237 Biological Activity #95: Regulation of viability or proliferation of immune cells (such as human eosinophil EOL-1 cells). 331 HMMAH60 1238 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 331 HMMAH60 1238 Biological Activity #65: Production of IL-10 and activation of T-cells. 332 HMQDF12 1239 Biological Activity #77: Production of MIP1alpha 332 HMQDF12 1239 Biological Activity #79: Production of TNF alpha by dendritic cells 333 HMQDT36 1240 Biological Activity #68: Production of IL-13 and activation of T-cells. 334 HMSBX80 1241 Biological Activity #101: Upregulation of CD71 and activation of T cells 335 HMSFS21 1242 Biological Activity #62: Production of ICAM-1 335 HMSFS21 1242 Biological Activity #72: Production of IL-6 336 HMSGB14 1243 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 337 HMSGU01 1244 Biological Activity #76: Production of MCP-1 338 HMSHM14 1245 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 338 HMSHM14 1245 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 338 HMSHM14 1245 Biological Activity #76: Production of MCP-1 339 HMSHS36 1246 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 339 HMSHS36 1246 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 339 HMSHS36 1246 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 339 HMSHS36 1246 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 340 HMSJM65 1247 Biological Activity #72: Production of IL-6 341 HMSJU68 1248 Biological Activity #12: Activation of Skeletal Muscle Cell PI3 Kinase Signaling Pathway 341 HMSJU68 1248 Biological Activity #38: Activation of transcription through NFKB response element in immune cells (such as EOL1 cells). 341 HMSJU68 1248 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 342 HMSKC04 1249 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 342 HMSKC04 1249 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 342 HMSKC04 1249 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 342 HMSKC04 1249 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 342 HMSKC04 1249 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 342 HMSKC04 1249 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 342 HMSKC04 1249 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 342 HMSKC04 1249 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 342 HMSKC04 1249 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 342 HMSKC04 1249 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 342 HMSKC04 1249 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 343 HMTAD67 1250 Biological Activity #65: Production of IL-10 and activation of T-cells. 344 HMUAP70 1251 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 345 HMVBN46 1252 Biological Activity #63: Production of IFNgamma using a T cells 346 HMWEB02 1253 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 347 HMWFO02 1254 Biological Activity #70: Production of IL-4 348 HMWFY10 1255 Biological Activity #60: Production of ICAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 348 HMWFY10 1255 Biological Activity #65: Production of IL-10 and activation of T-cells. 348 HMWFY10 1255 Biological Activity #75: Production of IL-8 by immune cells (such as the human EOL-1 eosinophil cells) 349 HMWGY65 1256 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 349 HMWGY65 1256 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 350 HNEAC05 1257 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 350 HNEAC05 1257 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 351 HNEEB45 1258 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 351 HNEEB45 1258 Biological Activity #38: Activation of transcription through NFKB response element in immune cells (such as EOL1 cells). 351 HNEEB45 1258 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 351 HNEEB45 1258 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 352 HNFFC43 1259 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 352 HNFFC43 1259 Biological Activity #82: Proliferation of immune cells (such as the HMC-1 human mast cell line) 352 HNFFC43 1259 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 352 HNFFC43 1259 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 353 HNFGF20 1260 Biological Activity #53: Bone marrow cell proliferation (fibronectin enhanced) 353 HNFGF20 1260 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 354 HNFJF07 1261 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 354 HNFJF07 1261 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 354 HNFJF07 1261 Biological Activity #93: Regulation of viability and proliferation of pancreatic beta cells. 354 HNFJF07 1261 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 355 HNFJH45 1262 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 355 HNFJH45 1262 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 355 HNFJH45 1262 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 356 HNGAK47 1263 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 356 HNGAK47 1263 Biological Activity #56: Endothelial Cell Apoptosis 357 HNGAP93 1264 Biological Activity #62: Production of ICAM-1 358 HNGBC07 1265 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 359 HNGBT31 1266 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 360 HNGDJ72 1267 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 360 HNGDJ72 1267 Biological Activity #60: Production of ICAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 360 HNGDJ72 1267 Biological Activity #72: Production of IL-6 360 HNGDJ72 1267 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 360 HNGDJ72 1267 Biological Activity #77: Production of MIP1alpha 360 HNGDJ72 1267 Biological Activity #79: Production of TNF alpha by dendritic cells 360 HNGDJ72 1267 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 360 HNGDJ72 1267 Biological Activity #101: Upregulation of CD71 and activation of T cells 360 HNGDJ72 1267 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 361 HNGDU40 1268 Biological Activity #98: Upregulation of CD152 and activation of T cells 362 HNGEG08 1269 Biological Activity #76: Production of MCP-1 363 HNGEO29 1270 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 364 HNGEP09 1271 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 364 HNGEP09 1271 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 365 HNGHR74 1272 Biological Activity #101: Upregulation of CD71 and activation of T cells 366 HNGIH43 1273 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 366 HNGIH43 1273 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 366 HNGIH43 1273 Biological Activity #76: Production of MCP-1 367 HNGIJ31 1274 Biological Activity #12: Activation of Skeletal Muscle Cell PI3 Kinase Signaling Pathway 367 HNGIJ31 1274 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 367 HNGIJ31 1274 Biological Activity #76: Production of MCP-1 367 HNGIJ31 1274 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 368 HNGIQ46 1275 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 369 HNGJE50 1276 Biological Activity #57: Insulin Secretion 369 HNGJE50 1276 Biological Activity #72: Production of IL-6 370 HNGJO57 1277 Biological Activity #79: Production of TNF alpha by dendritic cells 371 HNGJP69 1278 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 371 HNGJP69 1278 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 371 HNGJP69 1278 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 371 HNGJP69 1278 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 371 HNGJP69 1278 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 371 HNGJP69 1278 Biological Activity #49: Activation of transcription through serum response element in pre-adipocytes. 372 HNGJT54 1279 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 372 HNGJT54 1279 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 372 HNGJT54 1279 Biological Activity #76: Production of MCP-1 373 HNGOI12 1280 Biological Activity #65: Production of IL-10 and activation of T-cells. 373 HNGOI12 1280 Biological Activity #96: Stimulation of Calcium Flux in pancreatic beta cells. 374 HNGOM56 1281 Biological Activity #47: Activation of transcription through serum response element in immune cells (such as T-cells). 375 HNHAH01 1282 Biological Activity #62: Production of ICAM-1 376 HNHCX60 1283 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 376 HNHCX60 1283 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 377 HNHCY64 1284 Biological Activity #63: Production of IFNgamma using a T cells 378 HNHCY94 1285 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 379 HNHDW38 1286 Biological Activity #101: Upregulation of CD71 and activation of T cells 380 HNHDW42 1287 Biological Activity #72: Production of IL-6 380 HNHDW42 1287 Biological Activity #100: Upregulation of CD69 and activation of T cells 381 HNHED17 1288 Biological Activity #72: Production of IL-6 382 HNHEI42 1289 Biological Activity #59: Production of GM-CSF 383 HNHFO29 1290 Biological Activity #84: Regulation (inhibition or activation) of immune cell proliferation. 384 HNHFU32 1291 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 385 HNHOD46 1292 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 385 HNHOD46 1292 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #20: Activation of transcription through cAMP response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 385 HNHOD46 1292 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 385 HNHOD46 1292 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 385 HNHOD46 1292 Biological Activity #40: Activation of transcription through NFKB response element in immune cells (such as natural killer cells). 385 HNHOD46 1292 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 385 HNHOD46 1292 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #49: Activation of transcription through serum response element in pre-adipocytes. 385 HNHOD46 1292 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 385 HNHOD46 1292 Biological Activity #72: Production of IL-6 385 HNHOD46 1292 Biological Activity #77: Production of MIP1alpha 385 HNHOD46 1292 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 386 HNHOG73 1293 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 386 HNHOG73 1293 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 386 HNHOG73 1293 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 387 HNTBL27 1294 Biological Activity #65: Production of IL-10 and activation of T-cells. 387 HNTBL27 1294 Biological Activity #85: Regulation of apoptosis in pancreatic beta cells. 388 HNTCE26 1295 Biological Activity #62: Production of ICAM-1 388 HNTCE26 1295 Biological Activity #79: Production of TNF alpha by dendritic cells 388 HNTCE26 1295 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 388 HNTCE26 1295 Biological Activity #100: Upregulation of CD69 and activation of T cells 389 HNTNI01 1296 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 389 HNTNI01 1296 Biological Activity #25: Activation of transcription through GAS response element in immune cells (such as eosinophils). 389 HNTNI01 1296 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 389 HNTNI01 1296 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 389 HNTNI01 1296 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 389 HNTNI01 1296 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 389 HNTNI01 1296 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 389 HNTNI01 1296 Biological Activity #38: Activation of transcription through NFKB response element in immune cells (such as EOL1 cells). 389 HNTNI01 1296 Biological Activity #39: Activation of transcription through NFKB response element in immune cells (such as mast cells). 389 HNTNI01 1296 Biological Activity #46: Activation of transcription through serum response element in immune cells (such as T-cells). 389 HNTNI01 1296 Biological Activity #49: Activation of transcription through serum response element in pre-adipocytes. 389 HNTNI01 1296 Biological Activity #50: Activation of transcription through STAT6 response element in immune cells (such as mast cells). 389 HNTNI01 1296 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 389 HNTNI01 1296 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 389 HNTNI01 1296 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 390 HOAAC90 1297 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 390 HOAAC90 1297 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 391 HOACB38 1298 Biological Activity #72: Production of IL-6 392 HOCNF19 1299 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 392 HOCNF19 1299 Biological Activity #70: Production of IL-4 392 HOCNF19 1299 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 393 HODDN65 1300 Biological Activity #62: Production of ICAM-1 394 HODDN92 1301 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 394 HODDN92 1301 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 394 HODDN92 1301 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 394 HODDN92 1301 Biological Activity #72: Production of IL-6 394 HODDN92 1301 Biological Activity #76: Production of MCP-1 394 HODDN92 1301 Biological Activity #77: Production of MIP1alpha 394 HODDN92 1301 Biological Activity #90: Regulation of transcription through the FAS promoter element in hepatocytes 395 HODDO08 1302 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 396 HODDW40 1303 Biological Activity #77: Production of MIP1alpha 396 HODDW40 1303 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 397 HODFN71 1304 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 397 HODFN71 1304 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 397 HODFN71 1304 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 397 HODFN71 1304 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 397 HODFN71 1304 Biological Activity #46: Activation of transcription through serum response element in immune cells (such as T-cells). 397 HODFN71 1304 Biological Activity #47: Activation of transcription through serum response element in immune cells (such as T-cells). 397 HODFN71 1304 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 398 HODGE68 1305 Biological Activity #47: Activation of transcription through serum response element in immune cells (such as T-cells). 399 HOEBK34 1306 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 399 HOEBK34 1306 Biological Activity #76: Production of MCP-1 399 HOEBK34 1306 Biological Activity #98: Upregulation of CD152 and activation of T cells 399 HOEBK34 1306 Biological Activity #100: Upregulation of CD69 and activation of T cells 400 HOEBZ89 1307 Biological Activity #70: Production of IL-4 401 HOEDB32 1308 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 401 HOEDB32 1308 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 401 HOEDB32 1308 Biological Activity #77: Production of MIP1alpha 401 HOEDB32 1308 Biological Activity #79: Production of TNF alpha by dendritic cells 402 HOEDE28 1309 Biological Activity #79: Production of TNF alpha by dendritic cells 403 HOEDH84 1310 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 404 HOFMQ33 1311 Biological Activity #46: Activation of transcription through serum response element in immune cells (such as T-cells). 404 HOFMQ33 1311 Biological Activity #94: Regulation of viability and proliferation of pancreatic beta cells. 405 HOFMT75 1312 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 405 HOFMT75 1312 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 406 HOFNC14 1313 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 407 HOFND85 1314 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 408 HOFNY91 1315 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 408 HOFNY91 1315 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 409 HOFOC33 1316 Biological Activity #40: Activation of transcription through NFKB response element in immune cells (such as natural killer cells). 410 HOGCK20 1317 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 411 HOGCK63 1318 Biological Activity #62: Production of ICAM-1 412 HOGCS52 1319 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 413 HOHBB49 1320 Biological Activity #79: Production of TNF alpha by dendritic cells 414 HOHBC68 1321 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 415 HOHBY12 1322 Biological Activity #61: Production of ICAM-1 416 HOHCC74 1323 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 417 HOHCH55 1324 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 418 HOSDJ25 1325 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 418 HOSDJ25 1325 Biological Activity #61: Production of ICAM-1 418 HOSDJ25 1325 Biological Activity #85: Regulation of apoptosis in pancreatic beta cells. 419 HOSEG51 1326 Biological Activity #72: Production of IL-6 420 HOSEQ49 1327 Biological Activity #77: Production of MIP1alpha 421 HOSFD58 1328 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 422 HOUCQ17 1329 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 422 HOUCQ17 1329 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 422 HOUCQ17 1329 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 422 HOUCQ17 1329 Biological Activity #87: Regulation of proliferation and/or differentiation in immune cells (such as mast cells). 423 HOUDK26 1330 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 424 HOUGG12 1331 Biological Activity #77: Production of MIP1alpha 425 HOVCA92 1332 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 426 HPASA81 1333 Biological Activity #72: Production of IL-6 427 HPBCU51 1334 Biological Activity #59: Production of GM-CSF 427 HPBCU51 1334 Biological Activity #95: Regulation of viability or proliferation of immune cells (such as human eosinophil EOL-1 cells). 428 HPDDC77 1335 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 428 HPDDC77 1335 Biological Activity #69: Production of IL-2 and activation of T cells 429 HPDWP28 1336 Biological Activity #98: Upregulation of CD152 and activation of T cells 430 HPFCL43 1337 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 431 HPFDG48 1338 Biological Activity #50: Activation of transcription through STAT6 response element in immune cells (such as mast cells). 431 HPFDG48 1338 Biological Activity #79: Production of TNF alpha by dendritic cells 432 HPIAQ68 1339 Biological Activity #72: Production of IL-6 432 HPIAQ68 1339 Biological Activity #76: Production of MCP-1 433 HPIBO15 1340 Biological Activity #72: Production of IL-6 433 HPIBO15 1340 Biological Activity #93: Regulation of viability and proliferation of pancreatic beta cells. 434 HPJBK12 1341 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 434 HPJBK12 1341 Biological Activity #57: Insulin Secretion 434 HPJBK12 1341 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 435 HPJCL22 1342 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 435 HPJCL22 1342 Biological Activity #98: Upregulation of CD152 and activation of T cells 436 HPJCW04 1343 Biological Activity #79: Production of TNF alpha by dendritic cells 437 HPJEX20 1344 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 438 HPMAI22 1345 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 438 HPMAI22 1345 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 439 HPMFP40 1346 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 440 HPMGJ45 1347 Biological Activity #98: Upregulation of CD152 and activation of T cells 441 HPQAC69 1348 Biological Activity #98: Upregulation of CD152 and activation of T cells 442 HRPBC80 1349 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 442 HRPBC80 1349 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 442 HRPBC80 1349 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 442 HRPBC80 1349 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 442 HRPBC80 1349 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 442 HRPBC80 1349 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 442 HRPBC80 1349 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 443 HPRSB76 1350 Biological Activity #65: Production of IL-10 and activation of T-cells. 444 HPVAB94 1351 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 445 HPWAY46 1352 Biological Activity #22: Activation of transcription through CD28 response element in immune cells (such as T-cells). 445 HPWAY46 1352 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 445 HPWAY46 1352 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 445 HPWAY46 1352 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 445 HPWAY46 1352 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 446 HPWAZ95 1353 Biological Activity #65: Production of IL-10 and activation of T-cells. 447 HPWDJ42 1354 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 448 HPZAB47 1355 Biological Activity #3: Activation of transcription through NFKB response element in neuronal cells (such as SKNMC cells). 448 HPZAB47 1355 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 448 HPZAB47 1355 Biological Activity #98: Upregulation of CD152 and activation of T cells 449 HRAAB15 1356 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 449 HRAAB15 1356 Biological Activity #63: Production of IFNgamma using a T cells 450 HRABA80 1357 Biological Activity #6: Activation of Endothelial Cell ERK Signaling Pathway. 450 HRABA80 1357 Biological Activity #57: Insulin Secretion 450 HRABA80 1357 Biological Activity #98: Upregulation of CD152 and activation of T cells 451 HRACD15 1358 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 451 HRACD15 1358 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 451 HRACD15 1358 Biological Activity #89: Regulation of transcription of Malic Enzyme in hepatocytes 452 HRACD80 1359 Biological Activity #72: Production of IL-6 453 HRDDV47 1360 Biological Activity #101: Upregulation of CD71 and activation of T cells 454 HRDFD27 1361 Biological Activity #8: Activation of Endothelial Cell p38 or JNK Signaling Pathway. 454 HRDFD27 1361 Biological Activity #40: Activation of transcription through NFKB response element in immune cells (such as natural killer cells). 454 HRDFD27 1361 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 455 HRTAE58 1362 Biological Activity #79: Production of TNF alpha by dendritic cells 456 HSATR82 1363 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 457 HSAUK57 1364 Biological Activity #72: Production of IL-6 458 HSAUL82 1365 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 458 HSAUL82 1365 Biological Activity #79: Production of TNF alpha by dendritic cells 459 HSAVD46 1366 Biological Activity #65: Production of IL-10 and activation of T-cells. 460 HSAVH65 1367 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 461 HSAVK10 1368 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 461 HSAVK10 1368 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 461 HSAVK10 1368 Biological Activity #72: Production of IL-6 461 HSAVK10 1368 Biological Activity #77: Production of MIP1alpha 462 HSAWZ41 1369 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 462 HSAWZ41 1369 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 462 HSAWZ41 1369 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 462 HSAWZ41 1369 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 462 HSAWZ41 1369 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 462 HSAWZ41 1369 Biological Activity #38: Activation of transcription through NFKB response element in immune cells (such as EOL1 cells). 462 HSAWZ41 1369 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 462 HSAWZ41 1369 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 463 HSAXA83 1370 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 464 HSAYM40 1371 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 464 HSAYM40 1371 Biological Activity #72: Production of IL-6 465 HSDAJ46 1372 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 466 HSDEK49 1373 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 466 HSDEK49 1373 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 467 HSDER95 1374 Biological Activity #72: Production of IL-6 468 HSDEZ20 1375 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 469 HSDJA15 1376 Biological Activity #5: Activation of Adipocyte PI3 Kinase Signaling Pathway 469 HSDJA15 1376 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 469 HSDJA15 1376 Biological Activity #71: Production of IL-5 470 HSDSB09 1377 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 470 HSDSB09 1377 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 470 HSDSB09 1377 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 470 HSDSB09 1377 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 470 HSDSB09 1377 Biological Activity #39: Activation of transcription through NFKB response element in immune cells (such as mast cells). 470 HSDSB09 1377 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 470 HSDSB09 1377 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 470 HSDSB09 1377 Biological Activity #49: Activation of transcription through serum response element in pre-adipocytes. 470 HSDSB09 1377 Biological Activity #50: Activation of transcription through STAT6 response element in immune cells (such as mast cells). 470 HSDSB09 1377 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 470 HSDSB09 1377 Biological Activity #88: Regulation of transcription of Malic Enzyme in adipocytes 470 HSDSB09 1377 Biological Activity #92: Regulation of transcription via DMEF1 response element in adipocytes and pre-adipocytes 470 HSDSB09 1377 Biological Activity #96: Stimulation of Calcium Flux in pancreatic beta cells. 470 HSDSB09 1377 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 471 HSDSE75 1378 Biological Activity #58: Myoblast cell proliferation 471 HSDSE75 1378 Biological Activity #72: Production of IL-6 472 HSFAM31 1379 Biological Activity #65: Production of IL-10 and activation of T-cells. 473 HSHAX21 1380 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 473 HSHAX21 1380 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 473 HSHAX21 1380 Biological Activity #77: Production of MIP1alpha 473 HSHAX21 1380 Biological Activity #79: Production of TNF alpha by dendritic cells 474 HSIAS17 1381 Biological Activity #72: Production of IL-6 474 HSIAS17 1381 Biological Activity #79: Production of TNF alpha by dendritic cells 475 HSIDX71 1382 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 476 HSKDA27 1383 Biological Activity #59: Production of GM-CSF 476 HSKDA27 1383 Biological Activity #85: Regulation of apoptosis in pancreatic beta cells. 477 HSKHZ81 1384 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 477 HSKHZ81 1384 Biological Activity #62: Production of ICAM-1 478 HSLCQ82 1385 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 479 HSLJG37 1386 Biological Activity #59: Production of GM-CSF 480 HSNAB12 1387 Biological Activity #76: Production of MCP-1 481 HSODE04 1388 Biological Activity #63: Production of IFNgamma using a T cells 482 HSPBF70 1389 Biological Activity #98: Upregulation of CD152 and activation of T cells 483 HSQCM10 1390 Biological Activity #65: Production of IL-10 and activation of T-cells. 483 HSQCM10 1390 Biological Activity #95: Regulation of viability or proliferation of immune cells (such as human eosinophil EOL-1 cells). 484 HSSAJ29 1391 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 485 HSSDX51 1392 Biological Activity #72: Production of IL-6 486 HSSFT08 1393 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 487 HSSGD52 1394 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 487 HSSGD52 1394 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 487 HSSGD52 1394 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 487 HSSGD52 1394 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 488 HSSJC35 1395 Biological Activity #86: Regulation of apoptosis of immune cells (such as mast cells). 489 HSTBJ86 1396 Biological Activity #98: Upregulation of CD152 and activation of T cells 490 HSUBW09 1397 Biological Activity #90: Regulation of transcription through the FAS promoter element in hepatocytes 490 HSUBW09 1397 Biological Activity #98: Upregulation of CD152 and activation of T cells 491 HSVAM10 1398 Biological Activity #63: Production of IFNgamma using a T cells 492 HSVBU91 1399 Biological Activity #9: Activation of Hepatocyte ERK Signaling Pathway 492 HSVBU91 1399 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 492 HSVBU91 1399 Biological Activity #22: Activation of transcription through CD28 response element in immune cells (such as T-cells). 492 HSVBU91 1399 Biological Activity #57: Insulin Secretion 493 HSXCG83 1400 Biological Activity #72: Production of IL-6 494 HSXEC75 1401 Biological Activity #65: Production of IL-10 and activation of T-cells. 495 HSXEQ06 1402 Biological Activity #69: Production of IL-2 and activation of T cells 496 HSYAV50 1403 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 496 HSYAV50 1403 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 496 HSYAV50 1403 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 497 HSYAV66 1404 Biological Activity #72: Production of IL-6 498 HSYAZ50 1405 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 499 HSYAZ63 1406 Biological Activity #5: Activation of Adipocyte PI3 Kinase Signaling Pathway 499 HSYAZ63 1406 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 500 HSYBG37 1407 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 500 HSYBG37 1407 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 501 HSZAF47 1408 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 502 HT3SF53 1409 Biological Activity #72: Production of IL-6 503 HT5GJ57 1410 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 503 HT5GJ57 1410 Biological Activity #76: Production of MCP-1 504 HTADX17 1411 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 504 HTADX17 1411 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 504 HTADX17 1411 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 505 HTDAF28 1412 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 505 HTDAF28 1412 Biological Activity #98: Upregulation of CD152 and activation of T cells 505 HTDAF28 1412 Biological Activity #100: Upregulation of CD69 and activation of T cells 505 HTDAF28 1412 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 506 HTEAF65 1413 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 507 HTEBI28 1414 Biological Activity #71: Production of IL-5 507 HTEBI28 1414 Biological Activity #101: Upregulation of CD71 and activation of T cells 508 HTEDF80 1415 Biological Activity #98: Upregulation of CD152 and activation of T cells 509 HTEDY42 1416 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 509 HTEDY42 1416 Biological Activity #99: Upregulation of CD154 and activation of T cells 510 HTEFU65 1417 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 510 HTEFU65 1417 Biological Activity #58: Myoblast cell proliferation 510 HTEFU65 1417 Biological Activity #63: Production of IFNgamma using a T cells 510 HTEFU65 1417 Biological Activity #89: Regulation of transcription of Malic Enzyme in hepatocytes 510 HTEFU65 1417 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 511 HTEGI42 1418 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 512 HTEHR24 1419 Biological Activity #70: Production of IL-4 512 HTEHR24 1419 Biological Activity #98: Upregulation of CD152 and activation of T cells 513 HTEHU31 1420 Biological Activity #65: Production of IL-10 and activation of T-cells. 514 HTEHU93 1421 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 514 HTEHU93 1421 Biological Activity #65: Production of IL-10 and activation of T-cells. 515 HTEIP36 1422 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 515 HTEIP36 1422 Biological Activity #65: Production of IL-10 and activation of T-cells. 516 HTEIV80 1423 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 517 HTEJN13 1424 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 517 HTEJN13 1424 Biological Activity #100: Upregulation of CD69 and activation of T cells 518 HTELM16 1425 Biological Activity #47: Activation of transcription through serum response element in immune cells (such as T-cells). 518 HTELM16 1425 Biological Activity #77: Production of MIP1alpha 519 HTEPG70 1426 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 519 HTEPG70 1426 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 519 HTEPG70 1426 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 519 HTEPG70 1426 Biological Activity #37: Activation of transcription through NFKB response element in immune cells (such as basophils). 519 HTEPG70 1426 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 519 HTEPG70 1426 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 519 HTEPG70 1426 Biological Activity #49: Activation of transcription through serum response element in pre-adipocytes. 520 HTGAU75 1427 Biological Activity #101: Upregulation of CD71 and activation of T cells 521 HTGEP89 1428 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 522 HTHBG43 1429 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 522 HTHBG43 1429 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 523 HTHCA18 1430 Biological Activity #59: Production of GM-CSF 524 HTHDJ94 1431 Biological Activity #72: Production of IL-6 525 HTHDS25 1432 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 526 HTJMA95 1433 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 526 HTJMA95 1433 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 526 HTJMA95 1433 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 526 HTJMA95 1433 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 526 HTJMA95 1433 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 526 HTJMA95 1433 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 526 HTJMA95 1433 Biological Activity #65: Production of IL-10 and activation of T-cells. 527 HTJML75 1434 Biological Activity #59: Production of GM-CSF 528 HTLBE23 1435 Biological Activity #16: Activation of transcription through AP1 response element in immune cells (such as T-cells). 529 HTLFE42 1436 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 530 HTLFE57 1437 Biological Activity #62: Production of ICAM-1 531 HTLGE31 1438 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 532 HTLHY14 1439 Biological Activity #55: Calcium flux in immune cells (such as monocytes) 533 HTLIT32 1440 Biological Activity #70: Production of IL-4 534 HTLIV19 1441 Biological Activity #33: Activation of transcription through NFAT response element in immune cells (such as natural killer cells). 534 HTLIV19 1441 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 534 HTLIV19 1441 Biological Activity #51: Activation of transcription through STAT6 response element in immune cells (such as natural killer cells). 535 HTNBO91 1442 Biological Activity #61: Production of ICAM-1 536 HTOAK16 1443 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 536 HTOAK16 1443 Biological Activity #60: Production of ICAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 536 HTOAK16 1443 Biological Activity #67: Production of IL-13 536 HTOAK16 1443 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 536 HTOAK16 1443 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 537 HTODK73 1444 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 538 HTODO72 1445 Biological Activity #98: Upregulation of CD152 and activation of T cells 539 HTOGR42 1446 Biological Activity #7: Activation of Endothelial Cell JNK Signaling Pathway. 539 HTOGR42 1446 Biological Activity #11: Activation of Natural Killer Cell ERK Signaling Pathway. 539 HTOGR42 1446 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 540 HTOHD42 1447 Biological Activity #65: Production of IL-10 and activation of T-cells. 541 HTOHM15 1448 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 542 HTOHT18 1449 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 542 HTOHT18 1449 Biological Activity #79: Production of TNF alpha by dendritic cells 543 HTOIZ02 1450 Biological Activity #56: Endothelial Cell Apoptosis 543 HTOIZ02 1450 Biological Activity #72: Production of IL-6 544 HTOJA73 1451 Biological Activity #63: Production of IFNgamma using a T cells 545 HTOJK60 1452 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 546 HTPBW79 1453 Biological Activity #17: Activation of transcription through AP1 response element in immune cells (such as T-cells). 546 HTPBW79 1453 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 546 HTPBW79 1453 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 546 HTPBW79 1453 Biological Activity #34: Activation of transcription through NFAT response element in immune cells (such as T-cells). 546 HTPBW79 1453 Biological Activity #42: Activation of transcription through NFKB response element in immune cells (such as T-cells). 547 HTSEW17 1454 Biological Activity #2: Activation of transcription through NFKB response element in immune cells (such as B-cells). 547 HTSEW17 1454 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 548 HTTBI76 1455 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 548 HTTBI76 1455 Biological Activity #100: Upregulation of CD69 and activation of T cells 549 HTTDB46 1456 Biological Activity #2: Activation of transcription through NFKB response element in immune cells (such as B-cells). 549 HTTDB46 1456 Biological Activity #70: Production of IL-4 550 HTWCT03 1457 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 550 HTWCT03 1457 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 550 HTWCT03 1457 Biological Activity #60: Production of ICAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 550 HTWCT03 1457 Biological Activity #75: Production of IL-8 by immune cells (such as the human EOL-1 eosinophil cells) 550 HTWCT03 1457 Biological Activity #79: Production of TNF alpha by dendritic cells 550 HTWCT03 1457 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 550 HTWCT03 1457 Biological Activity #95: Regulation of viability or proliferation of immune cells (such as human eosinophil EOL-1 cells). 551 HTWDF76 1458 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 552 HTWJK32 1459 Biological Activity #76: Production of MCP-1 553 HTWKE60 1460 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 553 HTWKE60 1460 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 554 HTXCV12 1461 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 554 HTXCV12 1461 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 555 HTXDW56 1462 Biological Activity #3: Activation of transcription through NFKB response element in neuronal cells (such as SKNMC cells). 555 HTXDW56 1462 Biological Activity #27: Activation of transcription through GAS response element in immune cells (such as T-cells). 555 HTXDW56 1462 Biological Activity #28: Activation of transcription through GAS response element in immune cells (such as T-cells). 555 HTXDW56 1462 Biological Activity #35: Activation of transcription through NFAT response in immune cells (such as T-cells). 555 HTXDW56 1462 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 556 HTXFL30 1463 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 556 HTXFL30 1463 Biological Activity #79: Production of TNF alpha by dendritic cells 556 HTXFL30 1463 Biological Activity #87: Regulation of proliferation and/or differentiation in immune cells (such as mast cells). 557 HTXKP61 1464 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 558 HUDBZ89 1465 Biological Activity #18: Activation of transcription through cAMP response element (CRE) in pre-adipocytes. 558 HUDBZ89 1465 Biological Activity #59: Production of GM-CSF 559 HUFBY15 1466 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 560 HUFEF62 1467 Biological Activity #98: Upregulation of CD152 and activation of T cells 561 HUKAH51 1468 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 561 HUKAH51 1468 Biological Activity #83: Protection from Endothelial Cell Apoptosis. 562 HUKBT29 1469 Biological Activity #65: Production of IL-10 and activation of T-cells. 562 HUKBT29 1469 Biological Activity #70: Production of IL-4 562 HUKBT29 1469 Biological Activity #73: Production of IL6 by primary human aortic smooth muscle or normal human dermal fibroblast cells (without or with costimulation with TNFalpha). 562 HUKBT29 1469 Biological Activity #79: Production of TNF alpha by dendritic cells 563 HUSAT94 1470 Biological Activity #77: Production of MIP1alpha 564 HUSBA88 1471 Biological Activity #67: Production of IL-13 565 HUSIG64 1472 Biological Activity #15: Activation of transcription through AP1 response element in immune cells (such as T-cells). 566 HUSXS50 1473 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 566 HUSXS50 1473 Biological Activity #38: Activation of transcription through NFKB response element in immune cells (such as EOL1 cells). 566 HUSXS50 1473 Biological Activity #55: Calcium flux in immune cells (such as monocytes) 567 HWAAD63 1474 Biological Activity #62: Production of ICAM-1 567 HWAAD63 1474 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 567 HWAAD63 1474 Biological Activity #90: Regulation of transcription through the FAS promoter element in hepatocytes 568 HWABA81 1475 Biological Activity #62: Production of ICAM-1 568 HWABA81 1475 Biological Activity #98: Upregulation of CD152 and activation of T cells 569 HWABY10 1476 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 569 HWABY10 1476 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 569 HWABY10 1476 Biological Activity #72: Production of IL-6 570 HWADJ89 1477 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 570 HWADJ89 1477 Biological Activity #97: Stimulation of insulin secretion from pancreatic beta cells. 571 HWBAO62 1478 Biological Activity #23: Activation of transcription through CD28 response element in immune cells (such as T-cells). 572 HWBAR14 1479 Biological Activity #80: Production of TNF alpha by T cells 573 HWBAR88 1480 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils). 573 HWBAR88 1480 Biological Activity #75: Production of IL-8 by immune cells (such as the human EOL-1 eosinophil cells) 574 HWBCB89 1481 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 574 HWBCB89 1481 Biological Activity #41: Activation of transcription through NFKB response element in immune cells (such as T-cells). 574 HWBCB89 1481 Biological Activity #45: Activation of transcription through serum response element in immune cells (such as natural killer cells). 574 HWBCB89 1481 Biological Activity #62: Production of ICAM-1 575 HWBCP79 1482 Biological Activity #4: Activation of Adipocyte ERK Signaling Pathway 575 HWBCP79 1482 Biological Activity #65: Production of IL-10 and activation of T-cells. 576 HWBDP28 1483 Biological Activity #32: Activation of transcription through NFAT response element in immune cells (such as mast cells). 576 HWBDP28 1483 Biological Activity #62: Production of ICAM-1 577 HWBEM18 1484 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 578 HWBFE57 1485 Biological Activity #98: Upregulation of CD152 and activation of T cells 578 HWBFE57 1485 Biological Activity #101: Upregulation of CD71 and activation of T cells 579 HWDAC39 1486 Biological Activity #98: Upregulation of CD152 and activation of T cells 580 HWDAH38 1487 Biological Activity #48: Activation of transcription through serum response element in immune cells (such as T-cells). 581 HWHGP71 1488 Biological Activity #52: Activation of transcription through STAT6 response element in immune cells (such as T-cells). 582 HWHGQ49 1489 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 583 HWHGU54 1490 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 583 HWHGU54 1490 Biological Activity #74: Production of IL-8 by endothelial cells (such as Human Umbilical Cord Endothelial Cells). 584 HWHGZ51 1491 Biological Activity #29: Activation of transcription through GAS response element in immune cells (such as T-cells). 584 HWHGZ51 1491 Biological Activity #38: Activation of transcription through NFKB response element in immune cells (such as EOL1 cells). 584 HWHGZ51 1491 Biological Activity #76: Production of MCP-1 584 HWHGZ51 1491 Biological Activity #98: Upregulation of CD152 and activation of T cells 584 HWHGZ51 1491 Biological Activity #102: Upregulation of HLA-DR and activation of T cells 585 HWHHL34 1492 Biological Activity #19: Activation of transcription through cAMP response element in immune cells (such as T-cells). 585 HWHHL34 1492 Biological Activity #30: Activation of transcription through GATA-3 response element in immune cells (such as mast cells). 586 HWHQS55 1493 Biological Activity #68: Production of IL-13 and activation of T-cells. 587 HWLEV32 1494 Biological Activity #70: Production of IL-4 588 HWLIH65 1495 Biological Activity #14: Activation of T-Cell p38 or JNK Signaling Pathway. 588 HWLIH65 1495 Biological Activity #81: Production of VCAM in endothelial cells (such as human umbilical vein endothelial cells (HUVEC)) 589 HYAAJ71 1496 Biological Activity #61: Production of ICAM-1 590 HYBAR01 1497 Biological Activity #65: Production of IL-10 and activation of T-cells. 590 HYBAR01 1497 Biological Activity #76: Production of MCP-1 591 HYBBE75 1498 Biological Activity #65: Production of IL-10 and activation of T-cells. 592 HAPSA79 1499 Biological Activity #10: Activation of JNK Signaling Pathway in immune cells (such as eosinophils).

TABLE 1D.2 Biological Activity No. Biological Activity Exemplary Activity Assay 1 Activation of Assays for the activation of transcription through the EGR response element are well-known in the art and may be used or routinely transcription through the modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to EGR (Early Growth regulate EGR transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription through Response) element in the EGR response element that may be used or routinely modified to test EGR response element activity of polypeptides of the immune cells (such as B-cells). invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Richards JD, et al., J Immunol, 166(6): 3855-3864 (2001); Dinkel, A, et al., J Exp Med, 188(12): 2215-2224 (1998); and, Newton, JS, et al., Eur J Immunol 1996 Apr; 26(4): 811-816 (1996), the contents of each of which are herein incorporated by reference in its entirety. Immune cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary epithelial cells that may be used according to these assays include the Raji cell line. 2 Activation of Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely transcription through modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to NFKB response element regulate NFKB transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription in immune cells (such as B-cells). through the NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Gri G, et al., Biol Chem, 273(11): 6431-6438 (1998); Pyatt DW, et al., Cell Biol Toxicol 2000; 16(1): 41-51 (2000); Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Valle Blazquez et al, Immunology 90(3): 455-460 (1997); Aramburau et al., J Exp Med 82(3): 801-810 (1995); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. Immune cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary immune cells that may be used according to these assays include the Reh B-cell line. 3 Activation of Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely transcription through modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to NFKB response element regulate NFKB transcription factors and modulate expression of neuronal genes. Exemplary assays for transcription through the in neuronal cells (such as NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the SKNMC cells). invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Gill JS, et al., Neurobiol Dis, 7(4): 448-461 (2000); Tamatani M, et al., J Biol Chem, 274(13): 8531-8538 (1999); Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Valle Blazquez et al, Immunology 90(3): 455-460 (1997); Aramburau et al., J Exp Med 82(3): 801-810 (1995); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. Neuronal cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary neuronal cells that may be used according to these assays include the SKNMC neuronal cell line. 4 Activation of Adipocyte Kinase assay. Kinase assays, for example an Elk-1 kinase assay, for ERK signal transduction that regulate cell proliferation or ERK Signaling Pathway differentiation are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to promote or inhibit cell proliferation, activation, and differentiation. Exemplary assays for ERK kinase activity that may be used or routinely modified to test ERK kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Le Marchand-Brustel Y, Exp Clin Endocrinol Diabetes 107(2): 126-132 (1999); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Mouse adipocyte cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse adipocyte cells that may be used according to these assays include 3T3-L1 cells. 3T3-L1 is an adherent mouse preadipocyte cell line that is a continuous substrain of 3T3 fibroblast cells developed through clonal isolation and undergo a pre-adipocyte to adipose-like conversion under appropriate differentiation conditions known in the art. 5 Activation of Adipocyte Kinase assay. Kinase assays, for example an GSK-3 assays, for PI3 kinase signal transduction that regulate glucose metabolism and PI3 Kinase Signaling cell survival are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention Pathway (including antibodies and agonists or antagonists of the invention) to promote or inhibit glucose metabolism and cell survival. Exemplary assays for PI3 kinase activity that may be used or routinely modified to test PI3 kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Nikoulina et al., Diabetes 49(2): 263-271 (2000); and Schreyer et al., Diabetes 48(8): 1662-1666 (1999), the contents of each of which are herein incorporated by reference in its entirety. Mouse adipocyte cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse adipocyte cells that may be used according to these assays include 3T3-L1 cells. 3T3-L1 is an adherent mouse preadipocyte cell line that is a continuous substrain of 3T3 fibroblast cells developed through clonal isolation and undergo a pre-adipocyte to adipose-like conversion under appropriate differentiation conditions known in the art. 6 Activation of Endothelial Kinase assay. Kinase assays, for example an Elk-1 kinase assay, for ERK signal transduction that regulate cell proliferation or Cell ERK Signaling differentiation are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention Pathway. (including antibodies and agonists or antagonists of the invention) to promote or inhibit cell proliferation, activation, and differentiation. Exemplary assays for ERK kinase activity that may be used or routinely modified to test ERK kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Berra et al., Biochem Pharmacol 60(8): 1171-1178 (2000); Gupta et al., Exp Cell Res 247(2): 495-504 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Endothelial cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary endothelial cells that may be used according to these assays include human umbilical vein endothelial cells (HUVEC), which are endothelial cells which line venous blood vessels, and are involved in functions that include, but are not limited to, angiogenesis, vascular permeability, vascular tone, and immune cell extravasation. 7 Activation of Endothelial Kinase assay. JNK kinase assays for signal transduction that regulate cell proliferation, activation, or apoptosis are well known in the Cell JNK Signaling art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or Pathway. antagonists of the invention) to promote or inhibit cell proliferation, activation, and apoptosis. Exemplary assays for JNK kinase activity that may be used or routinely modified to test JNK kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Gupta et al., Exp Cell Res 247(2): 495-504 (1999); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Endothelial cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary endothelial cells that may be used according to these assays include human umbilical vein endothelial cells (HUVEC), which are endothelial cells which line venous blood vessels, and are involved in functions that include, but are not limited to, angiogenesis, vascular permeability, vascular tone, and immune cell extravasation. 8 Activation of Endothelial Kinase assay. JNK and p38 kinase assays for signal transduction that regulate cell proliferation, activation, or apoptosis are well Cell p38 or JNK known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and Signaling Pathway. agonists or antagonists of the invention) to promote or inhibit cell proliferation, activation, and apoptosis. Exemplary assays for JNK and p38 kinase activity that may be used or routinely modified to test JNK and p38 kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Gupta et al., Exp Cell Res 247(2): 495-504 (1999); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Endothelial cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary endothelial cells that may be used according to these assays include human umbilical vein endothelial cells (HUVEC), which are endothelial cells which line venous blood vessels, and are involved in functions that include, but are not limited to, angiogenesis, vascular permeability, vascular tone, and immune cell extravasation. 9 Activation of Hepatocyte Kinase assay. Kinase assays, for example an Elk-1 kinase assay, for ERK signal transduction that regulate cell proliferation or ERK Signaling Pathway differentiation are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to promote or inhibit cell proliferation, activation, and differentiation. Exemplary assays for ERK kinase activity that may be used or routinely modified to test ERK kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Rat liver hepatoma cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary rat liver hepatoma cells that may be used according to these assays include H4lle cells, which are known to respond to glucocorticoids, insulin, or cAMP derivatives. 10 Activation of JNK Kinase assay. JNK kinase assays for signal transduction that regulate cell proliferation, activation, or apoptosis are well known in the Signaling Pathway in art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or immune cells (such as antagonists of the invention) to promote or inhibit cell proliferation, activation, and apoptosis. Exemplary assays for JNK kinase eosinophils). activity that may be used or routinely modified to test JNK kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Gupta et al., Exp Cell Res 247(2): 495-504 (1999); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Exemplary cells that may be used according to these assays include eosinophils. Eosinophils are important in the late stage of allergic reactions; they are recruited to tissues and mediate the inflammatory response of late stage allergic reaction. Moreover, exemplary assays that may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to modulate signal transduction, cell proliferation, activation, or apoptosis in eosinophils include assays disclosed and/or cited in: Zhang JP, et al., “Role of caspases in dexamethasone- induced apoptosis and activation of c-Jun NH2-terminal kinase and p38 mitogen-activated protein kinase in human eosinophils” Clin Exp Immunol; Oct; 122(1): 20-7 (2000); Hebestreit H, et al., “Disruption of fas receptor signaling by nitric oxide in eosinophils” J Exp Med; Feb 2; 187(3): 415-25 (1998); J Allergy Clin Immunol 1999 Sep; 104(3 Pt 1): 565-74; and, Sousa AR, et al., “In vivo resistance to corticosteroids in bronchial asthma is associated with enhanced phosyphorylation of JUN N-terminal kinase and failure of prednisolone to inhibit JUN N-terminal kinase phosphorylation” J Allergy Clin Immunol; Sep; 104(3 Pt 1): 565-74 (1999); the contents of each of which are herein incorporated by reference in its entirety. 11 Activation of Natural Kinase assay. Kinase assays, for example an Elk-1 kinase assay, for ERK signal transduction that regulate cell proliferation or Killer Cell ERK Signaling differentiation are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention Pathway. (including antibodies and agonists or antagonists of the invention) to promote or inhibit cell proliferation, activation, and differentiation. Exemplary assays for ERK kinase activity that may be used or routinely modified to test ERK kinase-induced activity of polypeptides of the invention (including antibodies and agnonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Natural killer cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary natural killer cells that may be used according to these assays include the human natural killer cell lines (for example, NK-YT cells which have cytolytic and cytotoxic activity) or primary NK cells. 12 Activation of Skeletal Kinase assay. Kinase assays, for example an GSK-3 kinase assay, for PI3 kinase signal transduction that regulate glucose metabolism Muscle Cell PI3 Kinase and cell survival are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention Signaling Pathway (including antibodies and agonists or antagonists of the invention) to promote or inhibit glucose metabolism and cell survival. Exemplary assays for PI3 kinase activity that may be used or routinely modified to test PI3 kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Nikoulina et al., Diabetes 49(2): 263-271 (2000); and Schreyer et al., Diabetes 48(8): 1662-1666 (1999), the contents of each of which are herein incorporated by reference in its entirety. Rat myoblast cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary rat myoblast cells that may be used according to these assays include L6 cells. L6 is an adherent rat myoblast cell line, isolated from primary cultures of rat thigh muscle, that fuses to form multinucleated myotubes and striated fibers after culture in differentiation media. 13 Activation of Skeletal Kinase assay. Kinase assays, for example Elk-1 kinase assays, for ERK signal transduction that regulate cell proliferation or Muscle Cell ERK differentiation are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention Signaling Pathway (including antibodies and agonists or antagonists of the invention) to promote or inhibit cell proliferation, activation, and differentiation. Exemplary assays for ERK kinase activity that may be used or routinely modified to test ERK kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Le Marchand-Brustel Y, Exp Clin Endocrinol Diabetes 107(2): 126-132 (1999); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Rat myoblast cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary rat myoblast cells that may be used according to these assays include L6 cells. L6 is an adherent rat myoblast cell line, isolated from primary cultures of rat thigh muscle, that fuses to form multinucleated myotubes and striated fibers after culture in differentiation media. 14 Activation of T-Cell p38 Kinase assay. JNK and p38 kinase assays for signal transduction that regulate cell proliferation, activation, or apoptosis are well or JNK Signaling known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and Pathway. agonists or antagonists of the invention) to promote or inhibit immune cell (e.g. T-cell) proliferation, activation, and apoptosis. Exemplary assays for JNK and p38 kinase activity that may be used or routinely modified to test JNK and p38 kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Gupta et al., Exp Cell Res 247(2): 495-504 (1999); Kyriakis JM, Biochem Soc Symp 64: 29-48 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse T cells that may be used according to these assays include the CTLL cell line, which is an IL-2 dependent suspension-culture cell line with cytotoxic activity. 15 Activation of transcription Assays for the activation of transcription through the AP1 response element are known in the art and may be used or routinely through AP1 response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells modulate growth and other cell functions. Exemplary assays for transcription through the AP1 response element that may be used or (such as T-cells). routinely modified to test AP1-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1988); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Rellahan et al., J Biol Chem 272(49): 30806-30811 (1997); Chang et al., Mol Cell Biol 18(9): 4986-4993 (1998); and Fraser et al., Eur J Immunol 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. Mouse T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse T cells that may be used according to these assays include the HT2 cell line, which is an IL-2 dependent suspension culture cell line that also responds to IL-4. 16 Activation of transcription Assays for the activation of transcription through the AP1 response element are known in the art and may be used or routinely through AP1 response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells modulate growth and other cell functions. Exemplary assays for transcription through the AP1 response element that may be used or (such as T-cells). routinely modified to test AP1-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1988); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Rellahan et al., J Biol Chem 272(49): 30806-30811 (1997); Chang et al., Mol Cell Biol 18(9): 4986-4993 (1998); and Fraser et al., Eur J Immunol 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse T cells that may be used according to these assays include the CTLL cell line, which is an IL-2 dependent suspension-culture cell line with cytotoxic activity. 17 Activation of transcription Assays for the activation of transcription through the AP1 response element are well-known in the art and may be used or routinely through AP1 response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells modulate growth and other cell functions. Exemplary assays for transcription through the AP1 response element that may be used or (such as T-cells). routinely modified to test AP1-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1988); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Rellahan et al., J Biol Chem 272(49): 30806-30811 (1997); Chang et al., Mol Cell Biol 18(9): 4986-4993 (1998); and Fraser et al., Eur J Immunol 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the SUPT cell line, which is an IL-2 and IL-4 responsive suspension-culture cell line. 18 Activation of transcription Assays for the activation of transcription through the cAMP response element are well-known in the art and may be used or routinely through cAMP response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element (CRE) in pre- increase cAMP, regulate CREB transcription factors, and modulate expression of genes involved in a wide variety of cell functions. adipocytes. For example, a 3T3-L1/CRE reporter assay may be used to identify factors that activate the cAMP signaling pathway. CREB plays a major role in adipogenesis, and is involved in differentiation into adipocytes. CRE contains the binding sequence for the transcription factor CREB (CRE binding protein). Exemplary assays for transcription through the cAMP response element that may be used or routinely modified to test cAMP-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Reusch et al., Mol Cell Biol 20(3): 1008-1020 (2000); and Klemm et al., J Biol Chem 273: 917-923 (1998), the contents of each of which are herein incorporated by reference in its entirety. Pre-adipocytes that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary mouse adipocyte cells that may be used according to these assays include 3T3-L1 cells. 3T3-L1 is an adherent mouse preadipocyte cell line that is a continuous substrain of 3T3 fibroblast cells developed through clonal isolation and undergo a pre-adipocyte to adipose-like conversion under appropriate differentiation conditions known in the art. 19 Activation of transcription Assays for the activation of transcription through the cAMP response element are well-known in the art and may be used or routinely through cAMP response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells increase cAMP and regulate CREB transcription factors, and modulate expression of genes involved in a wide variety of cell functions. (such as T-cells). Exemplary assays for transcription through the cAMP response element that may be used or routinely modified to test cAMP-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Black et al., Virus Genes 15(2): 105-117 (1997); and Belkowski et al., J Immunol 161(2): 659-665 (1998), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse T cells that may be used according to these assays include the CTLL cell line, which is a suspension culture of IL-2 dependent cytotoxic T cells. 20 Activation of transcription Assays for the activation of transcription through the cAMP response element are well-known in the art and may be used or routinely through cAMP response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells increase cAMP, bind to CREB transcription factor, and modulate expression of genes involved in a wide variety of cell functions. (such as T-cells). Exemplary assays for transcription through the cAMP response element that may be used or routinely modified to test cAMP-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Black et al., Virus Genes 15(2): 105-117 (1997); and Belkowski et al., J Immunol 161(2): 659-665 (1998), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the JURKAT cell line, which is a suspension culture of leukemia cells that produce IL-2 when stimulated. 21 Activation of transcription Assays for the activation of transcription through the cAMP response element are well-known in the art and may be used or routinely through cAMP response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells increase cAMP, regulate CREB transcription factors, and modulate expression of genes involved in a wide variety of cell functions. (such as T-cells). Exemplary assays for transcription through the cAMP response element that may be used or routinely modified to test cAMP-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Black et al., Virus Genes 15(2): 105-117 (1997); and Belkowski et al., J Immunol 161(2): 659-665 (1998), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse T cells that may be used according to these assays include the HT2 cell line, which is a suspension culture of IL-2 dependent T cells that also respond to IL-4. 22 Activation of transcription Assays for the activation of transcription through the CD28 response element are well-known in the art and may be used or routinely through CD28 response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells stimulate IL-2 expression in T cells. Exemplary assays for transcription through the CD28 response element that may be used or (such as T-cells). routinely modified to test CD28-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); McGuire and Iacobelli, J Immunol 159(3): 1319-1327 (1997); Parra et al., J Immunol 166(4): 2437-2443 (2001); and Butscher et al., J Biol Chem 3(1): 552-560 (1998), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the JURKAT cell line, which is a suspension culture of leukemia cells that produce IL-2 when stimulated. 23 Activation of transcription Assays for the activation of transcription through the CD28 response element are well-known in the art and may be used or routinely through CD28 response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells stimulate IL-2 expression in T cells. Exemplary assays for transcription through the CD28 response element that may be used or (such as T-cells). routinely modified to test CD28-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); McGuire and Iacobelli, J Immunol 159(3): 1319-1327 (1997); Parra et al., J Immunol 166(4): 2437-2443 (2001); and Butscher et al., J Biol Chem 3(1): 552-560 (1998), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the SUPT cell line, which is a suspension culture of IL-2 and IL-4 responsive T cells. 24 Activation of transcription Assays for the activation of transcription through the Gamma Interferon Activation Site (GAS) response element are well-known in the through GAS response art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or element in epithelial cells antagonists of the invention) to regulate STAT transcription factors and modulate gene expression involved in a wide variety of cell (such as HELA cells). functions. Exemplary assays for transcription through the GAS response element that may be used or routinely modified to test GAS- response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: You M, et al, J Biol Chem, 272(37): 23376-23381(1997); Min W, et al., Circ Res, 83(8): 815-823 (1998); Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Matikainen et al., Blood 93(6): 1980-1991 (1999); and Henttinen et al., J Immunol 155(10): 4582-4587 (1995), the contents of each of which are herein incorporated by reference in its entirety. Epithelial cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary epithelial cells that may be used according to these assays include the HELA cell line. 25 Activation of transcription Assays for the activation of transcription through the Gamma Interferon Activation Site (GAS) response element are well-known in the through GAS response art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or element in immune cells antagonists of the invention) to modulate gene expression (commonly via STAT transcription factors) involved in a wide variety of cell (such as eosinophils). functions. Exemplary assays for transcription through the GAS response element that may be used or routinely modified to test GAS- response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Matikainen et al., Blood 93(6): 1980-1991 (1999); and Henttinen et al., J Immunol 155(10): 4582-4587 (1995); the contents of each of which are herein incorporated by reference in its entirety. Moreover, exemplary assays that may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to activate or inhibit activation of immune cells include assays disclosed and/or cited in: Mayumi M., “EoL-1, a human eosinophilic cell line” Leuk Lymphoma; Jun; 7(3): 243-50 (1992); Bhattacharya S, “Granulocyte macrophage colony- stimulating factor and interleukin-5 activate STAT5 and induce CIS1 mRNA in human peripheral blood eosinophils” Am J Respir Cell Mol Biol; Mar; 24(3): 312-6 (2001); and, Du J, et al., “Engagement of the CrkL adapter in interleukin-5 signaling in eosinophils” J Biol Chem; Oct 20; 275(42): 33167-75 (2000); the contents of each of which are herein incorporated by reference in its entirety. Exemplary cells that may be used according to these assays include eosinophils. Eosinophils are a type of immune cell important in the late stage of allergic reactions; they are recruited to tissues and mediate the inflammatory response of late stage allergic reaction. Increases in GAS mediated transcription in eosinophils is typically a result of STAT activation, normally a direct consequence of interleukin or other cytokine receptor stimulation (e.g. IL3, IL5 or GMCSF). 26 Activation of transcription Assays for the activation of transcription through the Gamma Interferon Activation Site (GAS) response element are well-known in the through GAS response art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or element in immune cells antagonists of the invention) to regulate STAT transcription factors and modulate gene expression involved in a wide variety of cell (such as monocytes). functions. Exemplary assays for transcription through the GAS response element that may be used or routinely modified to test GAS- response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Gustafson KS, et al., J Biol Chem, 271(33): 20035-20046 (1996); Eilers A, et al., Immunobiology, 193(2-4): 328-333 (1995); Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Matikainen et al., Blood 93(6): 1980-1991 (1999); and Henttinen et al., J Immunol 155(10): 4582-4587 (1995), the contents of each of which are herein incorporated by reference in its entirety. Exemplary immune cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary immune cells that may be used according to these assays include the U937 cell line, which is a monocytic cell line. 27 Activation of transcription Assays for the activation of transcription through the Gamma Interferon Activation Site (GAS) response element are well-known in the through GAS response art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or element in immune cells antagonists of the invention) to regulate STAT transcription factors and modulate gene expression involved in a wide variety of cell (such as T-cells). functions. Exemplary assays for transcription through the GAS response element that may be used or routinely modified to test GAS- response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Matikainen et al., Blood 93(6): 1980-1991 (1999); and Henttinen et al., J Immunol 155(10): 4582-4587 (1995), the contents of each of which are herein incorporated by reference in its entirety. Exemplary human T cells, such as the MOLT4 cell line, that may be used according to these assays are publicly available (e.g., through the ATCC). 28 Activation of transcription Assays for the activation of transcription through the Gamma Interferon Activation Site (GAS) response element are well-known in the through GAS response art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or element in immune cells antagonists of the invention) to regulate STAT transcription factors and modulate gene expression involved in a wide variety of cell (such as T-cells). functions. Exemplary assays for transcription through the GAS response element that may be used or routinely modified to test GAS- response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Matikainen et al., Blood 93(6): 1980-1991 (1999); and Henttinen et al., J Immunol 155(10): 4582-4587 (1995), the contents of each of which are herein incorporated by reference in its entirety. Exemplary human T cells, such as the SUPT cell line, that may be used according to these assays are publicly available (e.g., through the ATCC). 29 Activation of transcription Assays for the activation of transcription through the Gamma Interferon Activation Site (GAS) response element are well-known in the through GAS response art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or element in immune cells antagonists of the invention) to regulate STAT transcription factors and modulate gene expression involved in a wide variety of cell (such as T-cells). functions. Exemplary assays for transcription through the GAS response element that may be used or routinely modified to test GAS- response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Matikainen et al., Blood 93(6): 1980-1991 (1999); and Henttinen et al., J Immunol 155(10): 4582-4587 (1995), the contents of each of which are herein incorporated by reference in its entirety. Exemplary mouse T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary T cells that may be used according to these assays include the CTLL cell line, which is a suspension culture of IL-2 dependent cytotoxic T cells. 30 Activation of transcription This reporter assay measures activation of the GATA-3 signaling pathway in HMC-1 human mast cell line. Activation of GATA-3 in through GATA-3 mast cells has been linked to cytokine and chemokine production. Assays for the activation of transcription through the GATA3 response element in response element are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention immune cells (such as (including antibodies and agonists or antagonists of the invention) to regulate GATA3 transcription factors and modulate expression of mast cells). mast cell genes important for immune response development. Exemplary assays for transcription through the GATA3 response element that may be used or routinely modified to test GATA3-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Flavell et al., Cold Spring Harb Symp Quant Biol 64: 563-571 (1999); Rodriguez-Palmero et al., Eur J Immunol 29(12): 3914-3924 (1999); Zheng and Flavell, Cell 89(4): 587-596 (1997); and Henderson et al., Mol Cell Biol 14(6): 4286-4294 (1994), the contents of each of which are herein incorporated by reference in its entirety. Mast cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human mast cells that may be used according to these assays include the HMC-1 cell line, which is an immature human mast cell line established from the peripheral blood of a patient with mast cell leukemia, and exhibits many characteristics of immature mast cells. 31 Activation of transcription Assays for the activation of transcription through the GATA3 response element are well-known in the art and may be used or routinely through GATA-3 modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to response element in regulate GATA3 transcription factors and modulate expression of genes important for Th2 immune response development. Exemplary immune cells (such as T- assays for transcription through the GATA3 response element that may be used or routinely modified to test GATA3-response element cells). activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Flavell et al., Cold Spring Harb Symp Quant Biol 64: 563-571 (1999); Rodriguez-Palmero et al., Eur J Immunol 29(12): 3914-3924 (1999); Zheng and Flavell, Cell 89(4): 587-596 (1997); and Henderson et al., Mol Cell Biol 14(6): 4286-4294 (1994), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse T cells that may be used according to these assays include the HT2 cell line, which is a suspension culture of IL-2 dependent T cells that also respond to IL-4. 32 Activation of transcription This reporter assay measures activation of the NFAT signaling pathway in HMC-1 human mast cell line. Activation of NFAT in mast through NFAT response cells has been linked to cytokine and chemokine production. Assays for the activation of transcription through the Nuclear Factor of element in immune cells Activated T cells (NFAT) response element are well-known in the art and may be used or routinely modified to assess the ability of (such as mast cells). polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate NFAT transcription factors and modulate expression of genes involved in immunomodulatory functions. Exemplary assays for transcription through the NFAT response element that may be used or routinely modified to test NFAT-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); De Boer et al., Int J Biochem Cell Biol 31(10): 1221-1236 (1999); Ali et al., J Immunol 165(12): 7215-7223 (2000); Hutchinson and McCloskey, J Biol Chem 270(27): 16333-16338 (1995), and Turner et al., J Exp Med 188: 527-537 (1998), the contents of each of which are herein incorporated by reference in its entirety. Mast cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human mast cells that may be used according to these assays include the HMC-1 cell line, which is an immature human mast cell line established from the peripheral blood of a patient with mast cell leukemia, and exhibits many characteristics of immature mast cells. 33 Activation of transcription Assays for the activation of transcription through the Nuclear Factor of Activated T cells (NFAT) response element are well-known in through NFAT response the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists element in immune cells or antagonists of the invention) to regulate NFAT transcription factors and modulate expression of genes involved in (such as natural killer immunomodulatory functions. Exemplary assays for transcription through the NFAT response element that may be used or routinely cells). modified to test NFAT-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Aramburu et al., J Exp Med 182(3): 801-810 (1995); De Boer et al., Int J Biochem Cell Biol 31(10): 1221-1236 (1999); Fraser et al., Eur J Immunol 29(3): 838-844 (1999); and Yeseen et al., J Biol Chem 268(19): 14285-14293 (1993), the contents of each of which are herein incorporated by reference in its entirety. NK cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human NK cells that may be used according to these assays include the NK-YT cell line, which is a human natural killer cell line with cytolytic and cytotoxic activity. 34 Activation of transcription Assays for the activation of transcription through the Nuclear Factor of Activated T cells (NFAT) response element are well-known in through NFAT response the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists element in immune cells or antagonists of the invention) to regulate NFAT transcription factors and modulate expression of genes involved in (such as T-cells). immunomodulatory functions. Exemplary assays for transcription through the NFAT response element that may be used or routinely modified to test NFAT-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Serfling et al., Biochim Biophys Acta 1498(1): 1-18 (2000); De Boer et al., Int J Biochem Cell Biol 31(10): 1221-1236 (1999); Fraser et al., Eur J Immunol 29(3): 838-844 (1999); and Yeseen et al., J Biol Chem 268(19): 14285-14293 (1993), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the SUPT cell line, which is a suspension culture of IL-2 and IL-4 responsive T cells. 35 Activation of transcription Assays for the activation of transcription through the Nuclear Factor of Activated T cells (NFAT) response element are well-known in through NFAT response the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists in immune cells such as or antagonists of the invention) to regulate NFAT transcription factors and modulate expression of genes involved in T-cells). immunomodulatory functions. Exemplary assays for transcription through the NFAT response element that may be used or routinely modified to test NFAT-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Serfling et al., Biochim Biophys Acta 1498(1): 1-18 (2000); De Boer et al., Int J Biochem Cell Biol 31(10): 1221-1236 (1999); Fraser et al., Eur J Immunol 29(3): 838-844 (1999); and Yeseen et al., J Biol Chem 268(19): 14285-14293 (1993), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the JURKAT cell line, which is a suspension culture of leukemia cells that produce IL-2 when stimulated. 36 Activation of transcription Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely through NFKB response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in epithelial cells regulate NFKB transcription factors and modulate expression of epithelial genes. Exemplary assays for transcription through the (such as HELA cells). NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Kaltschmidt B, et al., Oncogene, 18(21): 3213-3225 (1999); Beetz A, et al., Int J Radiat Biol, 76(11): 1443-1453 (2000); Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Valle Blazquez et al, Immunology 90(3): 455-460 (1997); Aramburau et al., J Exp Med 82(3): 801-810 (1995); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. Epithelial cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary epithelial cells that may be used according to these assays include the HELA cell line. 37 Activation of transcription This reporter assay measures activation of the NFkB signaling pathway in Ku812 human basophil cell line. Assays for the activation of through NFKB response transcription through the NFKB response element are well-known in the art and may be used or routinely modified to assess the ability element in immune cells of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate NFKB transcription (such as basophils). factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription through the NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Marone et al, Int Arch Allergy Immunol 114(3): 207-17 (1997), the contents of each of which are herein incorporated by reference in its entirety. Basophils that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human basophil cell lines that may be used according to these assays include Ku812, originally established from a patient with chronic myelogenous leukemia. It is an immature prebasophilic cell line that can be induced to differentiate into mature basophils. 38 Activation of transcription Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely through the NFKB response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells regulate NFKB transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription (such as EOL1 cells). through the NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Valle Blazquez et al, Immunology 90(3): 455-460 (1997); Aramburau et al., J Exp Med 82(3): 801-810 (1995); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. For example, a reporter assay (which measures increases in transcription inducible from a NFkB responsive element in EOL-1 cells) may link the NFKB element to a reporter gene and binds to the NFKB transcription factor, which is upregulated by cytokines and other factors. Exemplary immune cells that may be used according to these assays include eosinophils such as the human EOL-1 cell line of eosinophils. Eosinophils are a type of immune cell important in the allergic responses; they are recruited to tissues and mediate the inflammatory response of late stage allergic reaction. Eol-1 is a human eosinophil cell line. 39 Activation of transcription This reporter assay measures activation of the NFkB signaling pathway in HMC-1 human mast cell line. Activation of NFkB in mast through NFKB response cells has been linked to production of certain cytokines, such as IL-6 and IL-9. Assays for the activation of transcription through the element in immune cells NFKB response element are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the (such as mast cells). invention (including antibodies and agonists or antagonists of the invention) to regulate NFKB transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription through the NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Stassen et al, J Immunol 166(7): 4391-8 (2001); and Marquardt and Walker, J Allergy Clin Immunol 105(3): 500-5 (2000), the contents of each of which are herein incorporated by reference in its entirety. Mast cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human mast cells that may be used according to these assays include the HMC-1 cell line, which is an immature human mast cell line established from the peripheral blood of a patient with mast cell leukemia, and exhibits many characteristics of immature mast cells. 40 Activation of transcription Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely through NFKB response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells regulate NFKB transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription (such as natural killer through the NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of cells). the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Valle Blazquez et al, Immunology 90(3): 455-460 (1997); Aramburau et al., J Exp Med 82(3): 801-810 (1995); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. NK cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary NK cells that may be used according to these assays include the NK-YT cell line, which is a human natural killer cell line with cytolytic and cytotoxic activity. 41 Activation of transcription Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely through NFKB response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells regulate NFKB transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription (such as T-cells). through the NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Black et al., Virus Gnes 15(2): 105-117 (1997); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. Exemplary human T cells, such as the MOLT4, that may be used according to these assays are publicly available (e.g., through the ATCC). 42 Activation of transcription Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely through NFKB response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells regulate NFKB transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription (such as T-cells). through the NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Black et al., Virus Gnes 15(2): 105-117 (1997); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the SUPT cell line, which is a suspension culture of IL-2 and IL-4 responsive T cells. 43 Activation of transcription Assays for the activation of transcription through the NFKB response element are well-known in the art and may be used or routinely through the NFKB response modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to element in immune cells regulate NFKB transcription factors and modulate expression of immunomodulatory genes. Exemplary assays for transcription (such as the Jurkat human through NFKB response element that may be used or routinely modified to test NFKB-response element activity of polypeptides of T cell line). the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Valle Blazquez et al, Immunology 90(3): 455-460 (1997); Aramburau et al., J Exp Med 82(3): 801-810 (1995); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the JURKAT cell line, which is a suspension culture of leukemia cells that produce IL-2 when stimulated. 44 Activation of transcription This assay uses a NFKB response element (which will bind NFKB transcription factors) linked to a reporter gene to measure NFKB through NFKB response mediated transcription in the human monocyte cell line U937. NFKB is upregulated by cytokines and other factors and NFKB element element in immune cells activation leads to expression of immunomodulatory genes. Activation of NFKB in monocytes can play a role in immune responses. (such as the U937 human Exemplary assays for transcription through the NFKB response element that may be used or routinely modified to test NFKB-response monocyte cell line). element activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Valle Blazquez et al, Immunology 90(3): 455-460 (1997); Aramburau et al., J Exp Med 82(3): 801-810 (1995); and Fraser et al., 29(3): 838-844 (1999), the contents of each of which are herein incorporated by reference in its entirety. Monocytic cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human monocyte cells that may be used according to these assays include the U937 cell line, which is cell line derived by Sundstrom and Nilsson in 1974 from malignant cells obtained from the pleural effusion of a patient with histiocytic lymphoma. 45 Activation of transcription Assays for the activation of transcription through the Serum Response Element (SRE) are well-known in the art and may be used or through serum response routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the element in immune cells invention) to regulate serum response factors and modulate the expression of genes involved in growth and upregulate the function of (such as natural killer growth-related genes in many cell types. Exemplary assays for transcription through the SRE that may be used or routinely modified cells). to test SRE activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Benson et al., J Immunol 153(9): 3862-3873 (1994); and Black et al., Virus Genes 12(2): 105-117 (1997), the content of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary T cells that may be used according to these assays include the NK-YT cell line, which is a human natural killer cell line with cytolytic and cytotoxic activity. 46 Activation of transcription Assays for the activation of transcription through the Serum Response Element (SRE) are well-known in the art and may be used or through serum response routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the element in immune cells invention) to bind the serum response factor and modulate the expression of genes involved in growth and upregulate the function of (such as T-cells). growth-related genes in many cell types. Exemplary assays for transcription through the SRE that may be used or routinely modified to test SRE activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Benson et al., J Immunol 153(9): 3862-3873 (1994); and Black et al., Virus Genes 12(2): 105-117 (1997), the content of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells, such as the MOLT4, that may be used according to these assays are publicly available (e.g., through the ATCC). 47 Activation of transcription Assays for the activation of transcription through the Serum Response Element (SRE) are well-known in the art and may be used or through serum response routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the element in immune cells invention) to regulate serum response factors and modulate the expression of genes involved in growth and upregulate the function of (such as T-cells). growth-related genes in many cell types. Exemplary assays for transcription through the SRE that may be used or routinely modified to test SRE activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Benson et al., J Immunol 153(9): 3862-3873 (1994); and Black et al., Virus Genes 12(2): 105-117 (1997), the content of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human T cells that may be used according to these assays include the JURKAT cell line, which is a suspension culture of leukemia cells that produce IL-2 when stimulated. 48 Activation of transcription Assays for the activation of transcription through the Serum Response Element (SRE) are well-known in the art and may be used or through serum response routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the element in immune cells invention) to regulate the serum response factors and modulate the expression of genes involved in growth. Exemplary assays for (such as T-cells). transcription through the SRE that may be used or routinely modified to test SRE activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); and Black et al., Virus Genes 12(2): 105-117 (1997), the content of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary mouse T cells that may be used according to these assays include the CTLL cell line, which is an IL-2 dependent suspension culture of T cells with cytotoxic activity. 49 Activation of transcription Assays for the activation of transcription through the Serum Response Element (SRE) are well-known in the art and may be used or through serum response routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the element in pre-adipocytes. invention) to regulate the serum response factors and modulate the expression of genes involved in growth. Exemplary assays for transcription through the SRE that may be used or routinely modified to test SRE activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); and Black et al., Virus Genes 12(2): 105-117 (1997), the content of each of which are herein incorporated by reference in its entirety. Pre- adipocytes that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary mouse adipocyte cells that may be used according to these assays include 3T3-L1 cells. 3T3-L1 is an adherent mouse preadipocyte cell line that is a continuous substrain of 3T3 fibroblast cells developed through clonal isolation and undergo a pre-adipocyte to adipose-like conversion under appropriate differentiation conditions known in the art. 50 Activation of transcription Assays for the activation of transcription through the Signal Transducers and Activators of Transcription (STAT6) response element in through STAT6 response immune cells (such as in the human HMC-1 mast cell line) are well-known in the art and may be used or routinely modified to assess element in immune cells the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate STAT6 (such as mast cells). transcription factors and modulate the expression of multiple genes. Exemplary assays for transcription through the STAT6 response element that may be used or routinely modified to test STAT6 response element activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Sherman, Immunol Rev 179: 48-56 (2001); Malaviya and Uckun, J Immunol 168: 421-426 (2002); Masuda et al., J Biol Chem 275(38): 29331-29337 (2000); and Masuda et al., J Biol Chem 276: 26107-26113 (2001), the contents of each of which are herein incorporated by reference in its entirety. Mast cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary human mast cells that may be used according to these assays include the HMC-1 cell line, which is an immature human mast cell line established from the peripheral blood of a patient with mast cell leukemia, and exhibits many characteristics of immature mast cells. 51 Activation of transcription Assays for the activation of transcription through the Signal Transducers and Activators of Transcription (STAT6) response element through STAT6 response are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including element in immune cells antibodies and agonists or antagonists of the invention) to regulate STAT6 transcription factors and modulate the expression of (such as natural killer multiple genes. Exemplary assays for transcription through the STAT6 response element that may be used or routinely modified to test cells). STAT6 response element activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Georas et al., Blood 92(12): 4529-4538 (1998); Moffatt et al., Transplantation 69(7): 1521-1523 (2000); Curiel et al., Eur J Immunol 27(8): 1982-1987 (1997); and Masuda et al., J Biol Chem 275(38): 29331-29337 (2000), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary rat natural killer cells that may be used according to these assays are publicly available (e.g., through the ATCC). 52 Activation of transcription Assays for the activation of transcription through the Signal Transducers and Activators of Transcription (STAT6) response element through STAT6 response are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including element in immune cells antibodies and agonists or antagonists of the invention) to regulate STAT6 transcription factors and modulate the expression of (such as T-cells). multiple genes. Exemplary assays for transcription through the STAT6 response element that may be used or routinely modified to test STAT6 response element activity of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Georas et al., Blood 92(12): 4529-4538 (1998); Moffatt et al., Transplantation 69(7): 1521-1523 (2000); Curiel et al., Eur J Immunol 27(8): 1982-1987 (1997); and Masuda et al., J Biol Chem 275(38): 29331-29337 (2000), the contents of each of which are herein incorporated by reference in its entirety. T cells that may be used according to these assays are publicly available (e.g., through the ATCC). Exemplary T cells that may be used according to these assays include the SUPT cell line, which is a suspension culture of IL-2 and IL-4 responsive T cells. 53 Bone marrow cell Assay for measuring regulation of proliferation of mouse bone marrow cells (in the presence or absence of exogenous Stem Cell proliferation (fibronectin Factor (SCF)) on a fibronectin extracellular matrix. Mouse bone marrow cells are plated onto 96-well fibronectin fragment coated enhanced) plates in 0.2 ml of serum-free medium. Secreted protein factors (test factors) are tested with appropriate negative controls in the presence and absence of SCF (5.0 ng/ml), where secreted test factor supernates represent 10% of the total assay volume. The cells are grown for 7 days. The number of proliferating cells within the wells is quantitated by measuring thymidine incorporation into cellular DNA. This and similar assays may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate proliferation of bone marrow cells. Interactions between adhesion receptors on progenitor cells and their extracellular matrix ligands are essential for the control of hematopoiesis in bone marrow stroma. These interactions may help retain CD34+ hematopoietic progenitor cells within the an appropriate bone marrow environment, and adhesive interactions can also provide important costimulatory signals. As the ability of stem cells to undergo self-renewal in vitro is dependent upon their interaction with the stromal cells and the extracellular matrix (ECM), this assay identify factors which integrate with the ECM environment and are important for stimulating stem cell self-renewal. 54 Calcium flux in Assays for measuring calcium flux are well-known in the art and may be used or routinely modified to assess the ability of chondrocytes polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mobilize calcium. Cells normally have very low concentrations of cytosolic calcium compared to much higher extracellular calcium. Extracellular factors can cause an influx of calcium, leading to activation of calcium responsive signaling pathways and alterations in cell functions. Exemplary assays that may be used or routinely modified to measure calcium flux in chondrocytes include assays disclosed in: Asada S, et al., Inflamm Res, 50(1): 19-23 (2001); Schwartz Z, et al., J Bone Miner Res, 6(7): 709-718 (1991); Iannotti JP, et al., J Bone Joint Surg Am, 67(1): 113-120 (1985); Sullivan E., et al., Methods Mol Biol 1999; 114: 125-133 (1999), the contents of each of which is herein incorporated by reference in its entirety. Cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary cells that may be used according to these assays include bovine chondrocytes. 55 Calcium flux in immune Assays for measuring calcium flux are well-known in the art and may be used or routinely modified to assess the ability of cells (such as monocytes) polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mobilize calcium. Cells normally have very low concentrations of cytosolic calcium compared to much higher extracellular calcium. Extracellular factors can cause an influx of calcium, leading to activation of calcium responsive signaling pathways and alterations in cell functions. Exemplary assays that may be used or routinely modified to measure calcium flux in immune cells (such as monocytes) include assays disclosed in: Chan, CC, et al., J Pharmacol Exp Ther, 269(3): 891-896 (1994); Andersson, K, et al., Cytokine, 12(12): 1784-1787 (2000); Scully, SP, et al., J Clin Invest, 74(2) 589-599 (1984); and, Sullivan, E, et al., Methods Mol Biol, 114: 125-133 (1999), the contents of each of which is herein incorporated by reference in its entirety. Cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary cells that may be used according to these assays include the THP-1 monocyte cell line. 56 Endothelial Cell Caspase Apoptosis. Assays for caspase apoptosis are well known in the art and may be used or routinely modified to assess the ability Apoptosis of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to promote caspase protease- mediated apoptosis. Induction of apoptosis in endothelial cells supporting the vasculature of tumors is associated with tumor regression due to loss of tumor blood supply. Exemplary assays for caspase apoptosis that may be used or routinely modified to test capase apoptosis activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Lee et al., FEBS Lett 485(2-3): 122-126 (2000); Nor et al., J Vasc Res 37(3): 209-218 (2000); and Karsan and Harlan, J Atheroscler Thromb 3(2): 75-80 (1996); the contents of each of which are herein incorporated by reference in its entirety. Endothelial cells that may be used according to these assays are publicly available (e.g., through commercial sources). Exemplary endothelial cells that may be used according to these assays include bovine aortic endothelial cells (bAEC), which are an example of endothelial cells which line blood vessels and are involved in functions that include, but are not limited to, angiogenesis, vascular permeability, vascular tone, and immune cell extravasation. 57 Insulin Secretion Assays for measuring secretion of insulin are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to stimulate insulin secretion. For example, insulin secretion is measured by FMAT using anti-rat insulin antibodies. Insulin secretion from pancreatic beta cells is upregulated by glucose and also by certain proteins/peptides, and disregulation is a key component in diabetes. Exemplary assays that may be used or routinely modified to test for stimulation of insulin secretion (from pancreatic cells) by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Shimizu, H., et al., Endocr J, 47(3): 261-9 (2000); Salapatek, A. M., et al., Mol Endocrinol, 13(8): 1305-17 (1999); Filipsson, K., et al., Ann N Y Acad Sci, 865: 441-4 (1998); Olson, L. K., et al., J Biol Chem, 271(28): 16544-52 (1996); and, Miraglia S et. al., Journal of Biomolecular Screening, 4: 193-204 (1999), the contents of each of which is herein incorporated by reference in its entirety. Pancreatic cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary pancreatic cells that may be used according to these assays include HITT15 Cells. HITT15 are an adherent epithelial cell line established from Syrian hamster islet cells transformed with SV40. These cells express glucagon, somatostatin, and glucocorticoid receptors. The cells secrete insulin, which is stimulated by glucose and glucagon and suppressed by somatostatin or glucocorticoids. ATTC# CRL-1777 Refs: Lord and Ashcroft. Biochem. J. 219: 547-551; Santerre et al. Proc. Natl. Acad. Sci. USA 78: 4339-4343, 1981. 58 Myoblast cell Assays for muscle cell proliferation are well known in the art and may be used or routinely modified to assess the ability of proliferation polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to stimulate or inhibit myoblast cell proliferation. Exemplary assays for myoblast cell proliferation that may be used or routinely modified to test activity of polypeptides and antibodies of the invention (including agonists or antagonists of the invention) include, for example, assays disclosed in: Soeta, C., et al. “Possible role for the c-ski gene in the proliferation of myogenic cells in regenerating skeletal muscles of rats” Dev Growth Differ Apr; 43(2): 155-64 (2001); Ewton DZ, et al., “IGF binding proteins-4, -5 and -6 may play specialized roles during L6 myoblast proliferation and differentiation” J Endocrinol Mar; 144(3): 539-53 (1995); and, Pampusch MS, et al., “Effect of transforming growth factor beta on proliferation of L6 and embryonic porcine myogenic cells” J Cell Physiol Jun; 143(3): 524-8 (1990); the contents of each of which are herein incorporated by reference in their entirety. Exemplary myoblast cells that may be used according to these assays include the rat myoblast L6 cell line. Rat myoblast L6 cells are an adherent rat myoblast cell line, isolated from primary cultures of rat thigh muscle, that fuse to form multinucleated myotubes and striated fibers after culture in differentiation media. 59 Production of GM-CSF GM-CSF FMAT. GM-CSF is expressed by activated T cells, macrophages, endothelial cells, and fibroblasts. GM-CSF regulates differentiation and proliferation of granulocytes-macrophage progenitors and enhances antimicrobial activity in neutrophils, monocytes and macrophage. Additionally, GM-CSF plays an important role in the differentiation of dendritic cells and monocytes, and increases antigen presentation. GM-CSF is considered to be a proinflammatory cytokine. Assays for immunomodulatory proteins that promote the production of GM-CSF are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation and modulate the growth and differentiation of leukocytes. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as GM-CSF, and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); and Ye et al., J Leukoc Biol (58(2): 225-233, the contents of each of which are herein incorporated by reference in its entirety. Natural killer cells that may be used according to these assays are publicly available (e.g., through the ATCC) or may be isolated using techniques disclosed herein or otherwise known in the art. Natural killer (NK) cells are large granular lymphocytes that have cytotoxic activity but do bind antigen. NK cells show antibody-independent killing of tumor cells and also recognize antibody bound on target cells, via NK Fc receptors, leading to cell-mediated cytotoxicity. 60 Production of ICAM in Endothelial cells, which are cells that line blood vessels, and are involved in functions that include, but are not limited to, angiogenesis, endothelial cells (such as vascular permeability, vascular tone, and immune cell extravasation. Exemplary endothelial cells that may be used in ICAM human umbilical vein production assays include human umbilical vein endothelial cells (HUVEC), and are available from commercial sources. The endothelial cells expression of ICAM (CD54), an integral membrane protein, can be upregulated by cytokines or other factors, and ICAM expression is (HUVEC)) important in mediating immune and endothelial cell interactions leading to immune and inflammatory responses. Assays for measuring expression of ICAM-1 are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate ICAM-1 expression. Exemplary assays that may be used or routinely modified to measure ICAM-1 expression include assays disclosed in: Rolfe BE, et al., Atherosclerosis, 149(1): 99-110 (2000); Panettieri RA Jr, et al., J Immunol, 154(5): 2358-2365 (1995); and, Grunstein MM, et al., Am J Physiol Lung Cell Mol Physiol, 278(6): L1154-L1163 (2000), the contents of each of which is herein incorporated by reference in its entirety. 61 Production of ICAM-1 Assays for measuring expression of ICAM-1 are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate ICAM-1 expression. Exemplary assays that may be used or routinely modified to measure ICAM-1 expression include assays disclosed in: Rolfe BE, et al., Atherosclerosis, 149(1): 99-110 (2000); Panettieri RA Jr, et al., J Immunol, 154(5): 2358-2365 (1995); and, Grunstein MM, et al., Am J Physiol Lung Cell Mol Physiol, 278(6): L1154-L1163 (2000), the contents of each of which is herein incorporated by reference in its entirety. Cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary cells that may be used according to these assays include Aortic Smooth Muscle Cells (AOSMC); such as bovine AOSMC. 62 Production of ICAM-1 Assays for measuring expression of ICAM-1 are well-known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate ICAM-1 expression. Exemplary assays that may be used or routinely modified to measure ICAM-1 expression include assays disclosed in: Takacs P, et al, FASEB J, 15(2): 279-281 (2001); and, Miyamoto K, et al., Am J Pathol, 156(5): 1733-1739 (2000), the contents of each of which is herein incorporated by reference in its entirety. Cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary cells that may be used according to these assays include microvascular endothelial cells (MVEC). 63 Production of IFNgamma IFNgamma FMAT. IFNg plays a central role in the immune system and is considered to be a proinflammatory cytokine. IFNg using a T cells promotes TH1 and inhibits TH2 differentiation; promotes IgG2a and inhibits IgE secretion; induces macrophage activation; and increases MHC expression. Assays for immunomodulatory proteins produced by T cells and NK cells that regulate a variety of inflammatory activities and inhibit TH2 helper cell functions are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, regulate inflammatory activities, modulate TH2 helper cell function, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as Interferon gamma (IFNg), and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Gonzalez et al., J Clin Lab Anal 8(5): 225-233 (1995); Billiau et al., Ann NY Acad Sci 856: 22-32 (1998); Boehm et al., Annu Rev Immunol 15: 749-795 (1997), and Rheumatology (Oxford) 38(3): 214-20 (1999), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T Cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 64 Production of IFNgamma IFNgamma FMAT. IFNg plays a central role in the immune system and is considered to be a proinflammatory cytokine. IFNg using Natural Killer cells promotes TH1 and inhibits TH2; promotes IgG2a and inhibits IgE; induces macrophage activation; and increases MHC expression. Assays for immunomodulatory proteins produced by T cells and NK cells that regulate a variety of inflammatory activities and inhibit TH2 helper cell functions are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, regulate inflammatory activities, modulate TH2 helper cell function, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as Interferon gamma (IFNg), and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Gonzalez et al., J Clin Lab Anal 8(5): 225-233 (1995); Billiau et al., Ann NY Acad Sci 856: 22-32 (1998); Boehm et al., Annu Rev Immunol 15: 749-795 (1997), and Rheumatology (Oxford) 38(3): 214-20 (1999), the contents of each of which are herein incorporated by reference in its entirety. Natural Killer (NK) cells that may be used according to these assays are publicly available (e.g., through the ATCC) or may be isolated using techniques disclosed herein or otherwise known in the art. Natural killer (NK) cells are large granular lymphocytes that have cytotoxic activity but do bind antigen. NK cells show antibody-independent killing of tumor cells and also recognize antibody bound on target cells, via NK Fc receptors, leading to cell-mediated cytotoxicity. 65 Production of IL-10 and Assays for production of IL-10 and activation of T-cells are well known in the art and may be used or routinely modified to assess the activation of T-cells. ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to stimulate or inhibit production of IL-10 and/or activation of T-cells. Exemplary assays that may be used or routinely modified to assess the ability of polypeptides and antibodies of the invention (including agonists or antagonists of the invention) to modulate IL-10 production and/or T-cell proliferation include, for example, assays such as disclosed and/or cited in: Robinson, DS, et al., “Th-2 cytokines in allergic disease” Br Med Bull; 56 (4): 956-968 (2000), and Cohn, et al., “T-helper type 2 cell-directed therapy for asthma” Pharmacology & Therapeutics; 88: 187-196 (2000); the contents of each of which are herein incorporated by reference in their entirety. Exemplary cells that may be used according to these assays include Th2 cells. IL10 secreted from Th2 cells may be measured as a marker of Th2 cell activation. Th2 cells are a class of T cells that secrete IL4, IL10, IL13, IL5 and IL6. Factors that induce differentiation and activation of Th2 cells play a major role in the initiation and pathogenesis of allergy and asthma. Primary T helper 2 cells are generated via in vitro culture under Th2 polarizing conditions using peripheral blood lymphocytes isolated from cord blood. 66 Production of IL-10 and IL-10 FMAT. Assays for immunomodulatory proteins produced by activated T cells, B cells, and monocytes that exhibit anti- downregulation of inflammatory activity and downregulate monocyte/macrophage function and expression of cytokines are well known in the art and may immune responses be used or routinely modified to assess the ability of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, regulate inflammatory activities, and modulate immune cell function and cytokine production. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as IL-10, and the downmodulation of immune responses. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); and Koning et al., Cytokine 9(6): 427-436 (1997), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 67 Production of IL-13 IL-13 FMAT. IL-13 enhances IgM, IgG, and IgE production and induces FcER1. IL-13 has anti-inflammatory activity on monocytes and macrophages. Assays for immunomodulatory proteins produced by T cells that inhibit activation and release of cytokines by macrophages are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, regulate cytokine release, stimulate immune cells through the binding of IL-13 and IL-4 receptors, and/or mediate humoral or cell-mediated immunity Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as IL-13, the inhibition of cytokines released by macrophages. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); and Ohshima et al., Blood 92(9): 3338-3345 (1998), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 68 Production of IL-13 and Assays for production of IL-13 and activation of T-cells are well known in the art and may be used or routinely modified to assess the activation of T-cells. ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to stimulate or inhibit production of IL-13 and/or activation of T-cells. Exemplary assays for IL-13 production that may be used or routinely modified to test activity of polypeptides and antibodies of the invention (including agonists or antagonists of the invention) include, for example, assays such as disclosed and/or cited in: Grunig, G, et al., “Requirement for IL-13 independently of IL-4 in Experimental asthma” Science; 282: 2261-2263 (1998), and Wills-Karp M, et al., “Interleukin-13: central mediator of allergic asthma” Science; 282: 2258-2261 (1998); the contents of each of which are herein incorporated by reference in their entirety. Exemplary cells that may be used according to these assays include Th2 cells. IL13, a Th2 type cytokine, is a potent stimulus for mucus production, airway hyper- responsiveness and allergic asthma. Th2 cells are a class of T cells that secrete IL4, IL10, IL13, IL5 and IL6. Factors that induce differentiation and activation of Th2 cells play a major role in the initiation and pathogenesis of allergy and asthma. Primary T helper 2 cells are generated in in vitro culture under Th2 polarizing conditions using peripheral blood lymphocytes isolated from cord blood. 69 Production of IL-2 and IL-2 FMAT. IL-2 is the principal T cell factor that allows T cell expansion and differentiation into effector cells. Assays for activation of T cells immunomodulatory proteins secreted by TH1 cells that promote T cell and NK cell growth and differentiation are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, promote immune cell growth and differentiation, and/or mediate humoral or cell-mediated immunity Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as IL-2, and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Laduda et al., Immunology 94(4): 496-502 (1998); and Powell et al., Immunol Rev 165: 287-300 (1998), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 70 Production of IL-4 IL-4 FMAT. Assays for immunomodulatory proteins secreted by TH2 cells that stimulate B cells, T cells, macrophages and mast cells and promote polarization of CD4+ cells into TH2 cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, stimulate immune cells, modulate immune cell polarization, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as IL-4, and the stimulation of immune cells, such as B cells, T cells, macrophages and mast cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Gonzalez et al., J Clin Lab Anal 8(5): 277-283 (1194); Yssel et al., Res Immunol 144(8): 610-616 (1993); Bagley et al., Nat Immunol 1(3): 257-261 (2000); and van der Graaff et al., Rheumatology (Oxford) 38(3): 214-220 (1999), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell- mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 71 Production of IL-5 IL-5 FMAT. Assays for immunomodulatory proteins secreted by TH2 cells, mast cells, basophils, and eosinophils that stimulate eosinophil function and B cell Ig production and promote polarization of CD4+ cells into TH2 cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, stimulate immune cell function, modulate B cell Ig production, modulate immune cell polarization, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as IL-5, and the stimulation of eosinophil function and B cell Ig production. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Ohshima et al., Blood 92(9): 3338-3345 (1998); Jung et al., Eur J Immunol 25(8): 2413-2416 (1995); Mori et al., J Allergy Clin Immunol 106(1 Pt 2): 558-564 (2000); and Koning et al., Cytokine 9(6): 427-436 (1997), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 72 Production of IL-6 IL-6 FMAT. IL-6 is produced by T cells and has strong effects on B cells. IL-6 participates in IL-4 induced IgE production and increases IgA production (IgA plays a role in mucosal immunity). IL-6 induces cytotoxic T cells. Deregulated expression of IL-6 has been linked to autoimmune disease, plasmacytomas, myelomas, and chronic hyperproliferative diseases. Assays for immunomodulatory and differentiation factor proteins produced by a large variety of cells where the expression level is strongly regulated by cytokines, growth factors, and hormones are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation and differentiation and modulate T cell proliferation and function. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as IL-6, and the stimulation and upregulation of T cell proliferation and functional activities. Such assays that may be used or routinely modified to test immunomodulatory and differentiation activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204(1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); and Verhasselt et al., J Immunol 158: 2919-2925 (1997), the contents of each of which are herein incorporated by reference in its entirety. Human dendritic cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human dendritic cells are antigen presenting cells in suspension culture, which, when activated by antigen and/or cytokines, initiate and upregulate T cell proliferation and functional activities. 73 Production of IL6 by Assay to measure regulation of production of Interleukin-6 (IL-6) by either human aortic smooth muscle cells or normal human dermal primary human aortic fibroblasts minus or plus costimulation with TNFalpha (TNFa). Human aortic smooth muscle cells or normal human dermal smooth muscle or normal fibroblasts may be obtained from commercial sources; these cells are important structural and functional components of blood vessels human dermal fibroblast and connective tissue, respectively. Interleukin-6 (IL-6) is a key molecule in chronic inflammation and has been implicated in the cells (without or with progression of arteriosclerosis, stroke, arthritis and other vascular and inflammatory diseases. Deregulated expression of IL-6 has been costimulation with linked to autoimmune disease, plasmacytomas, myelomas, and chronic hyperproliferative diseases. Assays for immunomodulatory and TNFalpha). differentiation factor proteins produced by a large variety of cells where the expression level is strongly regulated by cytokines, growth factors, and hormones are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation and production of IL-6. 74 Production of IL-8 by Assays measuring production of IL-8 are well known in the art and may be used or routinely modified to assess the ability of endothelial cells (such as polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate production and/or secretion Human Umbilical Cord of IL-8. For example, FMAT may be used or routinely modified to assess the ability of polypeptides of the invention (including Endothelial Cells). antibodies and agonists or antagonists of the invention) to regulate production and/or secretion of IL-8 from endothelial cells (such as human umbilical vein endothelial cells (HUVEC)). HUVECs are endothelial cells which line venous blood vessels, and are involved in functions that include, but are not limited to, angiogenesis, vascular permeability, vascular tone, and immune cell extravasation. Endothelial cells play a pivotal role in the initiation and perpetuation of inflammation and secretion of IL-8 may play an important role in recruitment and activation of immune cells such as neutrophils, macrophages, and lymphocytes. 75 Production of IL-8 by Assay that measures the production of the chemokine interleukin-8 (IL-8) from immune cells (such as the EOL-1 human eosinophil immune cells (such as the cell line) are well known in the art (for example, measurement of IL-8 production by FMAT) and may be used or routinely modified to human EOL-1 eosinophil assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to promote or cells) inhibit. Eosinophils are a type of immune cell important in allergic responses; they are recruited to tissues and mediate the inflammatory response of late stage allergic reaction. IL8 is a strong immunomodulator and may have a potential proinflammatory role in immunological diseases and disorders (such as allergy and asthma). 76 Production of MCP-1 MCP-1 FMAT. Assays for immunomodulatory proteins that are produced by a large variety of cells and act to induce chemotaxis and activation of monocytes and T cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, induce chemotaxis, and modulate immune cell activation. Exemplary assays that test for immunomodulatory proteins evaluate the production of cell surface markers, such as monocyte chemoattractant protein (MCP), and the activation of monocytes and T cells. Such assays that may be used or routinely modified to test immunomodulatory and differentiation activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204(1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Satthaporn and Eremin, J R Coll Surg Ednb 45(1): 9-19 (2001); and Verhasselt et al., J Immunol 158: 2919-2925 (1997), the contents of each of which are herein incorporated by reference in its entirety. Human dendritic cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human dendritic cells are antigen presenting cells in suspension culture, which, when activated by antigen and/or cytokines, initiate and upregulate T cell proliferation and functional activities. 77 Production of MIP1alpha MIP-1alpha FMAT. Assays for immunomodulatory proteins produced by activated dendritic cells that upregulate monocyte/macrophage and T cell chemotaxis are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, modulate chemotaxis, and modulate T cell differentiation. Exemplary assays that test for immunomodulatory proteins evaluate the production of chemokines, such as macrophage inflammatory protein 1 alpha (MIP-1a), and the activation of monocytes/macrophages and T cells. Such assays that may be used or routinely modified to test immunomodulatory and chemotaxis activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204(1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Satthaporn and Eremin, J R Coll Surg Ednb 45(1): 9-19 (2001); Drakes et al., Transp Immunol 8(1): 17-29 (2000); Verhasselt et al., J Immunol 158: 2919-2925 (1997); and Nardelli et al., J Leukoc Biol 65: 822-828 (1999), the contents of each of which are herein incorporated by reference in its entirety. Human dendritic cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human dendritic cells are antigen presenting cells in suspension culture, which, when activated by antigen and/or cytokines, initiate and upregulate T cell proliferation and functional activities. 78 Production of RANTES RANTES FMAT. Assays for immunomodulatory proteins that induce chemotaxis of T cells, monocytes, and eosinophils are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, induce chemotaxis, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines, such as RANTES, and the induction of chemotactic responses in immune cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000): Cocchi et al., Science 270(5243): 1811-1815 (1995); and Robinson et al., Clin Exp Immunol 101(3): 398-407 (1995), the contents of each of which are herein incorporated by reference in its entirety. Human immune cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. 79 Production of TNF alpha TNFa FMAT. Assays for immunomodulatory proteins produced by activated macrophages, T cells, fibroblasts, smooth muscle, and by dendritic cells other cell types that exert a wide variety of inflammatory and cytotoxic effects on a variety of cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, modulate inflammation and cytotoxicity. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines such as tumor necrosis factor alpha (TNFa), and the induction or inhibition of an inflammatory or cytotoxic response. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204(1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Verhasselt et al., Eur J Immunol 28(11): 3886-3890 (1198); Dahlen et al., J Immunol 160(7): 3585-3593 (1998); Verhasselt et al., J Immunol 158: 2919-2925 (1997); and Nardelli et al., J Leukoc Biol 65: 822-828 (1999), the contents of each of which are herein incorporated by reference in its entirety. Human dendritic cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human dendritic cells are antigen presenting cells in suspension culture, which, when activated by antigen and/or cytokines, initiate and upregulate T cell proliferation and functional activities. 80 Production of TNF alpha TNFa FMAT. Assays for immunomodulatory proteins produced by activated macrophages, T cells, fibroblasts, smooth muscle, and by T cells other cell types that exert a wide variety of inflammatory and cytotoxic effects on a variety of cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mediate immunomodulation, modulate inflammation and cytotoxicity, and mediate humoral and/or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the production of cytokines such as tumor necrosis factor alpha (TNFa), and the induction or inhibition of an inflammatory or cytotoxic response. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Verhasselt et al., Eur J Immunol 28(11): 3886-3890 (1198); Dahlen et al., J Immunol 160(7): 3585-3593 (1998); Verhasselt et al., J Immunol 158: 2919-2925 (1997); and Nardelli et al., J Leukoc Biol 65: 822-828 (1999), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 81 Production of VCAM in Assays for measuring expression of VCAM are well-known in the art and may be used or routinely modified to assess the ability of endothelial cells (such as polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate VCAM expression. For human umbilical vein example, FMAT may be used to measure the upregulation of cell surface VCAM-1 expression in endothelial cells. Endothelial cells endothelial cells are cells that line blood vessels, and are involved in functions that include, but are not limited to, angiogenesis, vascular permeability, (HUVEC)) vascular tone, and immune cell extravasation. Exemplary endothelial cells that may be used according to these assays include human umbilical vein endothelial cells (HUVEC), which are available from commercial sources. The expression of VCAM (CD106), a membrane-associated protein, can be upregulated by cytokines or other factors, and contributes to the extravasation of lymphocytes, leucocytes and other immune cells from blood vessels; thus VCAM expression plays a role in promoting immune and inflammatory responses. 82 Proliferation of immune Assays for the regulation (i.e. increases or decreases) of viability and proliferation of cells in vitro are well-known in the art and may cells (such as the HMC-1 be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of human mast cell line) the invention) to regulate viability and proliferation of eosinophil cells and cell lines. For example, the CellTiter-Gloo Luminescent Cell Viability Assay (PROMEGA ™ Corp., Madison, WI, USA) can be used to measure the number of viable cells in culture based on quantitation of the ATP present which signals the presence of metabolically active cells. Mast cells are found in connective and mucosal tissues throughout the body. Mast cell activation (via immunoglobulin E-antigen, promoted by T helper cell type 2 cytokines) is an important component of allergic disease. Dysregulation of mast cell apoptosis may play a role in allergic disease and mast cell tumor survival. Mast cell lines that may be used according to these assays are publicly available and/or may be routinely generated. Exemplary mast cells that may be used according to these assays include HMC-1, which is an immature human mast cell line established from the peripheral blood of a patient with mast cell leukemia, and exhibits many characteristics of immature mast cells. 83 Protection from Caspase Apoptosis Rescue. Assays for caspase apoptosis rescue are well known in the art and may be used or routinely modified to Endothelial Cell assess the ability of the polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to inhibit Apoptosis. caspase protease-mediated apoptosis. Exemplary assays for caspase apoptosis that may be used or routinely modified to test caspase apoptosis rescue of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in Romeo et al., Cardiovasc Res 45(3): 788-794 (2000); Messmer et al., Br J Pharmacol 127(7): 1633-1640 (1999); and J Atheroscler Thromb 3(2): 75-80 (1996); the contents of each of which are herein incorporated by reference in its entirety. Endothelial cells that may be used according to these assays are publicly available (e.g., through commercial sources). Exemplary endothelial cells that may be used according to these assays include bovine aortic endothelial cells (bAEC), which are an example of endothelial cells which line blood vessels and are involved in functions that include, but are not limited to, angiogenesis, vascular permeability, vascular tone, and immune cell extravasation. 84 Regulation (inhibition or The mixed lymphocyte reaction assay (MLR) (see e.g., Example: “Detection of Inhibition of a Mixed Lymphocyte Reaction” below) is activation) of immune cell a complex in vitro assay of T-cell responsiveness and immune cell activation. This assay is useful, for example, as an in vitro model of proliferation. allograft rejection and graft versus host disease. In this assays PBMCs from human donors are mixed, cultured, and monitored for thymidine incorporation (a measure of cell proliferation) to identify polypeptides of the invention (including antibodies and agonists or antagonists of the invention) that may activate or inhibit immune responses. 85 Regulation of apoptosis in Caspase Apoptosis. Assays for caspase apoptosis are well known in the art and may be used or routinely modified to assess the pancreatic beta cells. ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to promote caspase protease- mediated apoptosis. Apoptosis in pancreatic beta is associated with induction and progression of diabetes. Exemplary assays for caspase apoptosis that may be used or routinely modified to test capase apoptosis activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in: Loweth, AC, et al., FEBS Lett, 400(3): 285-8 (1997); Saini, KS, et al., Biochem Mol Biol Int, 39(6): 1229-36 (1996); Krautheim, A., et al., Br J Pharmacol, 129(4): 687-94 (2000); Chandra J, et al., Diabetes, 50 Suppl 1: S44-7 (2001); Suk K, et al., J Immunol, 166(7): 4481-9 (2001); Tejedo J, et al., FEBS Lett, 459(2): 238-43 (1999); Zhang, S., et al., FEBS Lett, 455(3): 315-20 (1999); Lee et al., FEBS Left 485(2-3): 122-126 (2000); Nor et al., J Vasc Res 37(3): 209-218 (2000); and Karsan and Harlan, J Atheroscler Thromb 3(2): 75-80 (1996); the contents of each of which are herein incorporated by reference in its entirety. Pancreatic cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary pancreatic cells that may be used according to these assays include RIN-m. RIN-m is a rat adherent pancreatic beta cell insulinoma cell line derived from a radiation induced transplantable rat islet cell tumor. The cells produce and secrete islet polypeptide hormones, and produce insulin, somatostatin, and possibly glucagon. ATTC: #CRL-2057 Chick et al. Proc. Natl. Acad. Sci. 1977 74: 628; AF et al. Proc. Natl. Acad. Sci. 1980 77: 3519. 86 Regulation of apoptosis of Caspase Apoptosis. Assays for caspase apoptosis are well known in the art and may be used or routinely modified to assess the ability immune cells (such as of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate caspase protease- mast cells). mediated apoptosis in immune cells (such as, for example, in mast cells). Mast cells are found in connective and mucosal tissues throughout the body, and their activation via immunoglobulin E-antigen, promoted by T helper cell type 2 cytokines, is an important component of allergic disease. Dysregulation of mast cell apoptosis may play a role in allergic disease and mast cell tumor survival. Exemplary assays for caspase apoptosis that may be used or routinely modified to test capase apoptosis activity induced by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in: Masuda A, et al., J Biol Chem, 276(28): 26107-26113 (2001); Yeatman CF 2nd, et al., J Exp Med, 192(8): 1093-1103 (2000); Lee et al., FEBS Lett 485(2-3): 122-126 (2000); Nor et al., J Vasc Res 37(3): 209-218 (2000); and Karsan and Harlan, J Atheroscler Thromb 3(2): 75-80 (1996); the contents of each of which are herein incorporated by reference in its entirety. Immune cells that may be used according to these assays are publicly available (e.g., through commercial sources). Exemplary immune cells that may be used according to these assays include mast cells such as the HMC human mast cell line. 87 Regulation of Kinase assays, for example an Elk-1 kinase assay for ERK signal transduction that regulates cell proliferation or differentiation, are proliferation and/or well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies differentiation in immune and agonists or antagonists of the invention) to promote or inhibit cell proliferation, activation, and differentiation. Exemplary assays cells (such as mast cells). for ERK kinase activity that may be used or routinely modified to test ERK kinase-induced activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include the assays disclosed in: Ali H, et al., J Immunol, 165(12): 7215-7223 (2000); Tam SY, et al., Blood, 90(5): 1807-1820 (1997); Forrer et al., Biol Chem 379(8-9): 1101-1110 (1998); Berra et al., Biochem Pharmacol 60(8): 1171-1178 (2000); Gupta et al., Exp Cell Res 247(2): 495-504 (1999); Chang and Karin, Nature 410(6824): 37-40 (2001); and Cobb MH, Prog Biophys Mol Biol 71(3-4): 479-500 (1999); the contents of each of which are herein incorporated by reference in its entirety. Exemplary immune cells that may be used according to these assays include human mast cells such as the HMC-1 cell line. 88 Regulation of Assays for the regulation of transcription of Malic Enzyme are well-known in the art and may be used or routinely modified to assess transcription of Malic the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate transcription Enzyme in adipocytes of Malic Enzyme, a key enzyme in lipogenesis. Malic enzyme is involved in lipogenesis and its expression is stimulated by insulin. ME promoter contains two direct repeat (DR1)-like elements MEp and MEd identified as putative PPAR response elements. ME promoter may also responds to AP1 and other transcription factors. Exemplary assays that may be used or routinely modified to test for regulation of transcription of Malic Enzyme (in adipoocytes) by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Streeper, R. S., et al., Mol Endocrinol, 12(11): 1778-91 (1998); Garcia- Jimenez, C., et al., Mol Endocrinol, 8(10): 1361-9 (1994); Barroso, I., et al., J Biol Chem, 274(25): 17997-8004 (1999); Ijpenberg, A., et al., J Biol Chem, 272(32): 20108-20117 (1997); Berger, et al., Gene 66: 1-10 (1988); and, Cullen, B., et al., Methods in Enzymol. 216: 362-368 (1992), the contents of each of which is herein incorporated by reference in its entirety. Hepatocytes that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary hepatocytes that may be used according to these assays includes the H4IIE rat liver hepatoma cell line. 89 Regulation of Assays for the regulation of transcription of Malic Enzyme are well-known in the art and may be used or routinely modified to assess transcription of Malic the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate transcription Enzyme in hepatocytes of Malic Enzyme, a key enzyme in lipogenesis. Malic enzyme is involved in lipogenesis and its expression is stimulated by insulin. ME promoter contains two direct repeat (DR1)-like elements MEp and MEd identified as putative PPAR response elements. ME promoter may also responds to AP1 and other transcription factors. Exemplary assays that may be used or routinely modified to test for regulation of transcription of Malic Enzyme (in hepatocytes) by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Streeper, R. S., et al., Mol Endocrinol, 12(11): 1778-91 (1998); Garcia- Jimenez, C., et al., Mol Endocrinol, 8(10): 1361-9 (1994); Barroso, I., et al., J Biol Chem, 274(25): 17997-8004 (1999); Ijpenberg, A., et al., J Biol Chem, 272(32): 20108-20117 (1997); Berger, et al., Gene 66: 1-10 (1988); and, Cullen, B., et al., Methods in Enzymol. 216: 362-368 (1992), the contents of each of which is herein incorporated by reference in its entirety. Hepatocytes that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary hepatocytes that may be used according to these assays includes the mouse 3T3-L1 cell line. 3T3-L1 is a mouse preadipocyte cell line (adherent). It is a continuous substrain of 3T3 fibroblasts developed through clonal isolation. Cells undergo a pre-adipocyte to adipose-like conversion under appropriate differentiation culture conditions. 90 Regulation of Assays for the regulation of transcription through the FAS promoter element are well-known in the art and may be used or routinely transcription through the modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to FAS promoter element in activate the FAS promoter element in a reporter construct and to regulate transcription of FAS, a key enzyme for lipogenesis. FAS hepatocytes promoter is regulated by many transcription factors including SREBP. Insulin increases FAS gene transcription in livers of diabetic mice. This stimulation of transcription is also somewhat glucose dependent. Exemplary assays that may be used or routinely modified to test for FAS promoter element activity (in hepatocytes) by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Xiong, S., et al., Proc Natl Acad Sci U.S.A., 97(8): 3948-53 (2000); Roder, K., et al., Eur J Biochem, 260(3): 743-51 (1999); Oskouian B, et al., Biochem J, 317 (Pt 1): 257-65 (1996); Berger, et al., Gene 66: 1-10 (1988); and, Cullen, B., et al., Methods in Enzymol. 216: 362-368 (1992), the contents of each of which is herein incorporated by reference in its entirety. Hepatocytes that may be used according to these assays, such as H4IIE cells, are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary hepatocytes that may be used according to these assays include rat liver hepatoma cell line(s) inducible with glucocorticoids, insulin, or cAMP derivatives. 91 Regulation of Assays for the regulation of transcription through the PEPCK promoter are well-known in the art and may be used or routinely transcription through the modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to PEPCK promoter in activate the PEPCK promoter in a reporter construct and regulate liver gluconeogenesis. Exemplary assays for regulation of hepatocytes transcription through the PEPCK promoter that may be used or routinely modified to test for PEPCK promoter activity (in hepatocytes) of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Berger et al., Gene 66: 1-10 (1998); Cullen and Malm, Methods in Enzymol 216: 362-368 (1992); Henthorn et al., Proc Natl Acad Sci USA 85: 6342-6346 (1988); Lochhead et al., Diabetes 49(6): 896-903 (2000); and Yeagley et al., J Biol Chem 275(23): 17814-17820 (2000), the contents of each of which is herein incorporated by reference in its entirety. Hepatocyte cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary liver hepatoma cells that may be used according to these assays include H4lle cells, which contain a tyrosine amino transferase that is inducible with glucocorticoids, insulin, or cAMP derivatives. 92 Regulation of Assays for the regulation of transcription through the DMEF1 response element are well-known in the art and may be used or routinely transcription via DMEF1 modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to response element in activate the DMEF1 response element in a reporter construct (such as that containing the GLUT4 promoter) and to regulate insulin adipocytes and pre- production. The DMEF1 response element is present in the GLUT4 promoter and binds to MEF2 transcription factor and another adipocytes transcription factor that is required for insulin regulation of Glut4 expression in skeletal muscle. GLUT4 is the primary insulin- responsive glucose transporter in fat and muscle tissue. Exemplary assays that may be used or routinely modified to test for DMEF1 response element activity (in adipocytes and pre-adipocytes) by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in Thai, M. V., et al., J Biol Chem, 273(23): 14285-92 (1998); Mora, S., et al., J Biol Chem, 275(21): 16323-8 (2000); Liu, M. L., et al., J Biol Chem, 269(45): 28514-21 (1994); “Identification of a 30-base pair regulatory element and novel DNA binding protein that regulates the human GLUT4 promoter in transgenic mice”, J Biol Chem. 2000 Aug 4; 275(31): 23666-73; Berger, et al., Gene 66: 1-10 (1988); and, Cullen, B., et al., Methods in Enzymol. 216: 362-368 (1992), the contents of each of which is herein incorporated by reference in its entirety. Adipocytes and pre-adipocytes that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary cells that may be used according to these assays include the mouse 3T3-L1 cell line which is an adherent mouse preadipocyte cell line. Mouse 3T3- L1 cells are a continuous substrain of 3T3 fibroblasts developed through clonal isolation. These cells undergo a pre-adipocyte to adipose-like conversion under appropriate differentiation culture conditions. 93 Regulation of viability Assays for the regulation of viability and proliferation of cells in vitro are well-known in the art and may be used or routinely modified and proliferation of to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate pancreatic beta cells. viability and proliferation of pancreatic beta cells. For example, the Cell Titer-Glo luminescent cell viability assay measures the number of viable cells in culture based on quantitation of the ATP present which signals the presence of metabolically active cells. Exemplary assays that may be used or routinely modified to test regulation of viability and proliferation of pancreatic beta cells by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Friedrichsen BN, et al., Mol Endocrinol, 15(1): 136-48 (2001); Huotari MA, et al., Endocrinology, 139(4): 1494-9 (1998); Hugl SR, et al., J Biol Chem 1998 Jul 10; 273(28): 17771-9 (1998), the contents of each of which is herein incorporated by reference in its entirety. Pancreatic cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary pancreatic cells that may be used according to these assays include rat INS-1 cells. INS-1 cells are a semi- adherent cell line established from cells isolated from an X-ray induced rat transplantable insulinoma. These cells retain characteristics typical of native pancreatic beta cells including glucose inducible insulin secretion. References: Asfari et al. Endocrinology 1992 130: 167. 94 Regulation of viability Assays for the regulation of viability and proliferation of cells in vitro are well-known in the art and may be used or routinely modified and proliferation of to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to regulate pancreatic beta cells. viability and proliferation of pancreatic beta cells. For example, the Cell Titer-Glo luminescent cell viability assay measures the number of viable cells in culture based on quantitation of the ATP present which signals the presence of metabolically active cells. Exemplary assays that may be used or routinely modified to test regulation of viability and proliferation of pancreatic beta cells by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Ohtani KI, et al., Endocrinology, 139(1): 172-8 (1998); Krautheim A, et al, Exp Clin Endocrinol Diabetes, 107 (1): 29-34 (1999), the contents of each of which is herein incorporated by reference in its entirety. Pancreatic cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary pancreatic cells that may be used according to these assays include HITT15 Cells. HITT15 are an adherent epithelial cell line established from Syrian hamster islet cells transformed with SV40. These cells express glucagon, somatostatin, and glucocorticoid receptors. The cells secrete insulin, which is stimulated by glucose and glucagon and suppressed by somatostatin or glucocorticoids. ATTC# CRL-1777 Refs: Lord and Ashcroft. Biochem. J. 219: 547-551; Santerre et al. Proc. Natl. Acad. Sci. USA 78: 4339-4343, 1981. 95 Regulation of viability or Assays for the regulation (i.e. increases or decreases) of viability and proliferation of cells in vitro are well-known in the art and may proliferation of immune be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of cells (such as human the invention) to regulate viability and proliferation of eosinophil cells and cell lines. For example, the CellTiter-Gloo Luminescent eosinophil EOL-1 cells). Cell Viability Assay (PROMEGA ™ Corp., Madison, WI, USA) can be used to measure the number of viable cells in culture based on quantitation of the ATP present which signals the presence of metabolically active cells. Eosinophils are a type of immune cell important in allergic responses; they are recruited to tissues and mediate the inflammatory response of late stage allergic reaction. Eosinophil cell lines that may be used according to these assays are publicly available and/or may be routinely generated. Exemplary eosinophil cells that may be used according to these assays include EOL-1 Cells. 96 Stimulation of Calcium Assays for measuring calcium flux are well-known in the art and may be used or routinely modified to assess the ability of Flux in pancreatic beta polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to mobilize calcium. For example, the cells. FLPR assay may be used to measure influx of calcium. Cells normally have very low concentrations of cytosolic calcium compared to much higher extracellular calcium. Extracellular factors can cause an influx of calcium, leading to activation of calcium responsive signaling pathways and alterations in cell functions. Exemplary assays that may be used or routinely modified to measure calcium flux by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Satin LS, et al., Endocrinology, 136(10): 4589-601 (1995); Mogami H, et al., Endocrinology, 136(7): 2960-6 (1995); Richardson SB, et al., Biochem J, 288 (Pt 3): 847-51 (1992); and, Meats, JE, et al., Cell Calcium 1989 Nov-Dec; 10(8): 535-41 (1989), the contents of each of which is herein incorporated by reference in its entirety. Pancreatic cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary pancreatic cells that may be used according to these assays include HITT15 Cells. HITT15 are an adherent epithelial cell line established from Syrian hamster islet cells transformed with SV40. These cells express glucagon, somatostatin, and glucocorticoid receptors. The cells secrete insulin, which is stimulated by glucose and glucagon and suppressed by somatostatin or glucocorticoids. ATTC# CRL-1777 Refs: Lord and Ashcroft. Biochem. J. 219: 547-551; Santerre et al. Proc. Natl. Acad. Sci. USA 78: 4339-4343, 1981. 97 Stimulation of insulin Assays for measuring secretion of insulin are well-known in the art and may be used or routinely modified to assess the ability of secretion from pancreatic polypeptides of the invention (including antibodies and agonists or antagonists of the insecretion from pancreaticvention) to stimulate insulin secretion. For beta cells. example, insulin secretion is measured by FMAT using anti-rat insulin antibodies. Insulin secretion from pancreatic beta cells is upregulated by glucose and also by certain proteins/peptides, and disregulation is a key component in diabetes. Exemplary assays that may be used or routinely modified to test for stimulation of insulin secretion (from pancreatic cells) by polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include assays disclosed in: Ahren, B., et al., Am J Physiol, 277(4 Pt 2): R959-66 (1999); Li, M., et al., Endocrinology, 138(9): 3735-40 (1997); Kim, K. H., et al., FEBS Left, 377(2): 237-9 (1995); and, Miraglia S et. al., Journal of Biomolecular Screening, 4: 193-204 (1999), the contents of each of which is herein incorporated by reference in its entirety. Pancreatic cells that may be used according to these assays are publicly available (e.g., through the ATCC) and/or may be routinely generated. Exemplary pancreatic cells that may be used according to these assays include rat INS-1 cells. INS-1 cells are a semi-adherent cell line established from cells isolated from an X-ray induced rat transplantable insulinoma. These cells retain characteristics typical of native pancreatic beta cells including glucose inducible insulin secretion. References: Asfari et al. Endocrinology 1992 130: 167. 98 Upregulation of CD152 CD152 FMAT. CD152 (a.k.a. CTLA-4) expression is restricted to activated T cells. CD152 is a negative regulator of T cell and activation of T cells proliferation. Reduced CD152 expression has been linked to hyperproliferative and autoimmune diseases. Overexpression of CD152 may lead to impaired immunoresponses. Assays for immunomodulatory proteins important in the maintenance of T cell homeostasis and expressed almost exclusively on CD4+ and CD8+ T cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to modulate the activation of T cells, maintain T cell homeostasis, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the upregulation of cell surface markers, such as CD152, and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include, for example, the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); McCoy et al, Immunol Cell Biol 77(1): 1-10 (1999); Oostervegal et al., Curr Opin Immunol 11(3): 294-300 (1999); and Saito T, Curr Opin Immunol 10(3): 313-321 (1998), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T Cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell- mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 99 Upregulation of CD154 CD154 FMAT. CD154 (a.k.a., CD40L) expression is induced following activation of T cells. Interaction between CD154 and CD40 and activation of T cells on B cells is required for correct antibody class switching and germinal center formation. Mutations in CD154 are linked to immunodeficiencies and increased susceptibility to infections. Assays for immunomodulatory proteins important for antibody class switching and TH1 function and expressed on activated T helper lymphocytes are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to modulate the activation of T cells, modulate antibody class switching, mediate TH1 function, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the upregulation of cell surface markers, such as CD154, and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include, for example, the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Mackey et al., J Leukoc Biol 63(4): 418:428 (1998); and Skov et al., 164(7): 3500-3505 (2000), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T Cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 100 Upregulation of CD69 CD69 FMAT. CD69 is an activation marker that is expressed on activated T cells, B cells, and NK cells. CD69 is not expressed on and activation of T cells resting T cells, B cells, or NK cells. CD69 has been found to be associated with inflammation. Assays for immunomodulatory proteins expressed in T cells, B cells, and leukocytes are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to modulate the activation of T cells, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the upregulation of cell surface markers, such as CD69, and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include, for example, the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Ferenczi et al., J Autoimmun 14(1): 63-78 (200); Werfel et al., Allergy 52(4): 465-469 (1997); Taylor-Fishwick and Siegel, Eur J Immunol 25(12): 3215-3221 (1995); and Afetra et al., Ann Rheum Dis 52(6): 457-460 (1993), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T Cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 101 Upregulation of CD71 CD71 FMAT. CD71 is the transferrin receptor. Transferrin is a major iron carrying protein that is essential for cell proliferation. and activation of T cells CD71 is expressed predominantly on cells that are actively proliferating. Assays for immunomodulatory proteins expressed on activated T cells, B cells, and most proliferating cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to modulate the activation of T cells, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the upregulation of cell surface markers, such as CD71, and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include, for example, the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); and Afetra et al., Ann Rheum Dis 52(6): 457-460 (1993), the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T Cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors. 102 Upregulation of HLA-DR HLA-DR FMAT. MHC class II is essential for correct presentation of antigen to CD4+ T cells. Deregulation of MHC class II has been and activation of T cells associated with autoimmune diseases (e.g., diabetes, rheumatoid arthritis, systemic lupus erythematosis, and multiple sclerosis). Assays for immunomodulatory proteins expressed on MHC class II expressing T cells and antigen presenting cells are well known in the art and may be used or routinely modified to assess the ability of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) to modulate the activation of T cells, and/or mediate humoral or cell-mediated immunity. Exemplary assays that test for immunomodulatory proteins evaluate the upregulation of MHC class II products, such as HLA-DR antigens, and the activation of T cells. Such assays that may be used or routinely modified to test immunomodulatory activity of polypeptides of the invention (including antibodies and agonists or antagonists of the invention) include, for example, the assays disclosed in Miraglia et al., J Biomolecular Screening 4: 193-204 (1999); Rowland et al., “Lymphocytes: a practical approach” Chapter 6: 138-160 (2000); Lamour et al., Clin Exp Immunol 89(2): 217-222 (1992); Hurme and Sihvola, Immunol Lett 20(3): 217-222 (1989); Gansbacher and Zier, Cell Immunol 117(1): 22-34 (1988); and Itoh et al., J Histochem Cytochem 40(11): 1675-1683, the contents of each of which are herein incorporated by reference in its entirety. Human T cells that may be used according to these assays may be isolated using techniques disclosed herein or otherwise known in the art. Human T cells are primary human lymphocytes that mature in the thymus and express a T Cell receptor and CD3, CD4, or CD8. These cells mediate humoral or cell-mediated immunity and may be preactivated to enhance responsiveness to immunomodulatory factors.

Table 1E: Polynucleotides encoding polypeptides of the present invention can be used in assays to test for one or more biological activities. One such biological activity which may be tested includes the ability of polynucleotides and polypeptides of the invention to stimulate up-regulation or down-regulation of expression of particular genes and proteins. Hence, if polynucleotides and polypeptides of the present invention exhibit activity in altering particular gene and protein expression patterns, it is likely that these polynucleotides and polypeptides of the present invention may be involved in, or capable of effecting changes in, diseases associated with the altered gene and protein expression profiles. Hence, polynucleotides, polypeptides, or antibodies of the present invention could be used to treat said associated diseases.

TaqMan® assays may be performed to assess the ability of polynucleotides (and polypeptides they encode) to alter the expression pattern of particular “target” genes. TaqMan® reactions are performed to evaluate the ability of a test agent to induce or repress expression of specific genes in different cell types. TaqMan® gene expression quantification assays (“TaqMan® assays”) are well known to, and routinely performed by, those of ordinary skill in the art. TaqMan® assays are performed in a two step reverse transcription/polymerase chain reaction (RT-PCR). In the first (RT) step, cDNA is reverse transcribed from total RNA samples using random hexamer primers. In the second (PCR) step, PCR products are synthesized from the cDNA using gene specific primers.

To quantify gene expression the Taqman® PCR reaction exploits the 5′ nuclease activity of AmpliTaq Gold® DNA Polymerase to cleave a Taqman® probe (distinct from the primers) during PCR. The Taqman® probe contains a reporter dye at the 5′-end of the probe and a quencher dye at the 3′ end of the probe. When the probe is intact, the proximity of the reporter dye to the quencher dye results in suppression of the reporter fluorescence. During PCR, if the target of interest is present, the probe specifically anneals between the forward and reverse primer sites. AmpliTaq Fold DNA Polymerase then cleaves the probe between the reporter and quencher when the probe hybridizes to the target, resulting in increased fluorescence of the reporter (see FIG. 2). Accumulation of PCR products is detected directly by monitoring the increase in fluorescence of the reporter dye.

After the probe fragments are displaced from the target, polymerization of the strand continues. The 3′-end of the probe is blocked to prevent extension of the probe during PCR. This process occurs in every cycle and does not interfere with the exponential accumulation of product. The increase in fluorescence signal is detected only if the target sequence is complementary to the probe and is amplified during PCR. Because of these requirements, any nonspecific amplification is not detected.

For test sample preparation, vector controls or constructs containing the coding sequence for the gene of interest are transfected into cells, such as for example 293T cells, and supernatants collected after 48 hours. For cell treatment and RNA isolation, multiple primary human cells or human cell lines are used; such cells may include but are not limited to, Normal Human Dermal Fibroblasts, Aortic Smooth Muscle, Human Umbilical Vein Endothelial Cells, HepG2, Daudi, Jurkat, U937, Caco, and THP-1 cell lines. Cells are plated in growth media and growth is arrested by culturing without media change for 3 days, or by switching cells to low serum media and incubating overnight. Cells are treated for 1, 6, or 24 hours with either vector control supernatant or sample supernatant (or purified/partially purified protein preparations in buffer). Total RNA is isolated; for example, by using Trizol extraction or by using the Ambion RNAqueous™-4PCR RNA isolation system. Expression levels of multiple genes are analyzed using TAQMAN, and expression in the test sample is compared to control vector samples to identify genes induced or repressed. Each of the above described techniques are well known to, and routinely performed by, those of ordinary skill in the art.

Table 1E indicates particular disease classes and preferred indications for which polynucleotides, polypeptides, or antibodies of the present invention may be used in detecting, diagnosing, preventing, treating and/or ameliorating said diseases and disorders based on “target” gene expression patterns which may be up- or down-regulated by polynucleotides (and the encoded polypeptides) corresponding to each indicated cDNA Clone ID (shown in Table 1E, Column 2).

Thus, in preferred embodiments, the present invention encompasses a method of detecting, diagnosing, preventing, treating, and/or ameliorating a disease or disorder listed in the “Disease Class” and/or “Preferred Indication” columns of Table 1E; comprising administering to a patient in which such detection, diagnosis, prevention, or treatment is desired a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof) in an amount effective to detect, diagnose, prevent, treat, or ameliorate the disease or disorder. The first and second columns of Table 1D.1 show the “Gene No.” and “cDNA Clone ID No.”, respectively, indicating certain nucleic acids and proteins (or antibodies against the same) of the invention (including polynucleotide, polypeptide, and antibody fragments or variants thereof) that may be used in detecting, diagnosing, preventing, treating, or ameliorating the disease(s) or disorder(s) indicated in the corresponding row in the “Disease Class” or “Preferred Indication” Columns of Table 1E.

In another embodiment, the present invention also encompasses methods of detecting, diagnosing, preventing, treating, or ameliorating a disease or disorder listed in the “Disease Class” or “Preferred Indication” Columns of Table 1E; comprising administering to a patient combinations of the proteins, nucleic acids, or antibodies of the invention (or fragments or variants thereof), sharing similar indications as shown in the corresponding rows in the “Disease Class” or “Preferred Indication” Columns of Table 1E.

The “Disease Class” Column of Table 1E provides a categorized descriptive heading for diseases, disorders, and/or conditions (more fully described below) that may be detected, diagnosed, prevented, treated, or ameliorated by a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof).

The “Preferred Indication” Column of Table 1E describes diseases, disorders, and/or conditions that may be detected, diagnosed, prevented, treated, or ameliorated by a protein, nucleic acid, or antibody of the invention (or fragment or variant thereof).

The “Cell Line” and “Exemplary Targets” Columns of Table 1E indicate particular cell lines and target genes, respectively, which may show altered gene expression patterns (i.e., up- or down-regulation of the indicated target gene) in Taqman assays, performed as described above, utilizing polynucleotides of the cDNA Clone ID shown in the corresponding row. Alteration of expression patterns of the indicated “Exemplary Target” genes is correlated with a particular “Disease Class” and/or “Preferred Indication” as shown in the corresponding row under the respective column headings.

The “Exemplary Accessions” Column indicates GenBank Accessions (available online through the National Center for Biotechnology Information (NCBI)) which correspond to the “Exemplary Targets” shown in the adjacent row.

The recitation of “Cancer” in the “Disease Class” Column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof) may be used for example, to detect, diagnose, prevent, treat, and/or ameliorate neoplastic diseases and/or disorders (e.g., leukemias, cancers, etc., as described below under “Hyperproliferative Disorders”).

The recitation of “Immune” in the “Disease Class” column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof), may be used for example, to detect, diagnose, prevent, treat, and/or ameliorate diseases and/or disorders relating to neoplastic diseases (e.g., as described below under “Hyperproliferative Disorders”), blood disorders (e.g., as described below under “Immune Activity” “Cardiovascular Disorders” and/or “Blood-Related Disorders”), and infections (e.g., as described below under “Infectious Disease”).

The recitation of “Angiogenesis” in the “Disease Class” column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof), may be used for example, to detect, diagnose, treat, prevent, and/or ameliorate diseases and/or disorders relating to neoplastic diseases (e.g., as described below under “Hyperproliferative Disorders”), diseases and/or disorders of the cardiovascular system (e.g., as described below under “Cardiovascular Disorders”), diseases and/or disorders involving cellular and genetic abnormalities (e.g., as described below under “Diseases at the Cellular Level”), diseases and/or disorders involving angiogenesis (e.g., as described below under “Anti-Angiogenesis Activity”), to promote or inhibit cell or tissue regeneration (e.g., as described below under “Regeneration”), or to promote wound healing (e.g., as described below under “Wound Healing and Epithelial Cell Proliferation”).

The recitation of “Diabetes” in the “Disease Class” column indicates that the corresponding nucleic acid and protein, or antibody against the same, of the invention (or fragment or variant thereof), may be used for example, to detect, diagnose, treat, prevent, and/or ameliorate diabetes (including diabetes mellitus types I and II), as well as diseases and/or disorders associated with, or consequential to, diabetes (e.g. as described below under “Endocrine Disorders,” “Renal Disorders,” and “Gastrointestinal Disorders”).

TABLE 1E Gene Exemplary No. CDNA Clone Disease Class Preferred Indications Cell Line Targets Exemplary Accessions 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as AOSMC CIS3 gb|AB006967|AB006967 described herein under the heading “Immune Activity” and/or “Blood- GATA1 gb|X17254|HSERYF1 Related Disorders” (particularly including, but not limited to, immune IL1B gb|X02532|HSIL1BR disorders involving muscle tissues and the cardiovascular system (e.g. heart, lungs, circulatory system)). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving muscle tissue or the cardiovascular system). (AOSMC cells are human aortic smooth muscle cells). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as Caco-2 TNF gb|AJ270944|HSA27094 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving the cells of the gastrointestinal tract). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the gastrointestinal tract). (The Caco-2 cell line is a human colorectal adenocarcinoma cell line available through the ATCC ™ as cell line number HTB-37). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as HEK293 GATA3 gb|X55037|HSGATA3 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). (The 293 cell line is a human embryonal kidney epithelial cell line available through the ATCC ™ as cell line number CRL-1573). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as HUVEC CD30 gb|X12705|HSBCDFIA described herein under the heading “Immune Activity” and/or “Blood- HLA-c gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune IL5 disorders involving endothelial cells). Highly preferred embodiments of TNF the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving endothelial cells). (HUVEC cells are human umbilical vein endothelial cells). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as Jurkat Rag1 gb|M29474|HUMRAG1 described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Jurkat cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number TIB-152). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as Liver LTBR gb|AK027080|AK027080 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving cells of the hepatic system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the hepatic system). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as SK-N-MC CIS3 gb|AB006967|AB006967 described herein under the heading “Immune Activity” and/or “Blood- neuroblastoma GATA1 gb|X17254|HSERYF1 Related Disorders” (particularly including, but not limited to, immune HLA-c disorders involving the central nervous system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the central nervous system). (The SK-N-MC neuroblastoma cell line is a cell line derived from human brain tissue and is available through the ATCC ™ as cell line number HTB-10). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as T-cell-Mar. 31, 2000 CD40 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- Granzyme B gb|J04071|HUMCSE Related Disorders” (particularly including, but not limited to, immune disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). 15 HAGDG59 Immune Highly preferred indications include immunological disorders such as U937 CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as Caco-2 CCR4 gb|AB023888|AB023888 described herein under the heading “Immune Activity” and/or “Blood- CIS3 gb|AB006967|AB006967 Related Disorders” (particularly including, but not limited to, immune ICAM gb|X06990|HSICAM1 disorders involving the cells of the gastrointestinal tract). Highly VCAM gb|A30922|A30922 preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the gastrointestinal tract). (The Caco-2 cell line is a human colorectal adenocarcinoma cell line available through the ATCC ™ as cell line number HTB-37). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as Daudi Rag1 gb|M29474|HUMRAG1 described herein under the heading “Immune Activity” and/or “Blood- Rag2 gb|AY011962|AY011962 Related Disorders” (particularly including, but not limited to, immune disorders involving the B-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving B-cells). (The Daudi cell line is a human B lymphoblast cell line available through the ATCC ™ as cell line number CCL-213). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as HUVEC CD25 gb|X03137|HSIL2RG7 described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving endothelial cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving endothelial cells). (HUVEC cells are human umbilical vein endothelial cells). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as Jurkat CD28 gb|AF222342|AF222342 described herein under the heading “Immune Activity” and/or “Blood- IL2 gb|X61155|HSARTIL2 Related Disorders” (particularly including, but not limited to, immune VCAM gb|A30922|A30922 disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Jurkat cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number TIB-152). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as Liver CCR4 gb|AB023888|AB023888 described herein under the heading “Immune Activity” and/or “Blood- CD28 gb|AF222342|AF222342 Related Disorders” (particularly including, but not limited to, immune CXCR3 gb|Z79783|HSCKRL2 disorders involving cells of the hepatic system). Highly preferred Rag2 gb|AY011962|AY011962 embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the hepatic system). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as NHDF CIS3 gb|AB006967|AB006967 described herein under the heading “Immune Activity” and/or “Blood- Rag1 gb|M29474|HUMRAG1 Related Disorders” (particularly including, but not limited to, immune disorders involving the skin). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the skin). (NHDF cells are normal human dermal fibroblasts). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as THP1 CD28 gb|AF222342|AF222342 described herein under the heading “Immune Activity” and/or “Blood- CIS3 gb|AB006967|AB006967 Related Disorders” (particularly including, but not limited to, immune CXCR3 gb|Z79783|HSCKRL2 disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The THP1 cell line is a human monocyte cell line available through the ATCC ™ as cell line number TIB-202). 80 HCHNF25 Immune Highly preferred indications include immunological disorders such as U937 TNF gb|AJ270944|HSA27094 described herein under the heading “Immune Activity” and/or “Blood- VCAM gb|A30922|A30922 Related Disorders” (particularly including, but not limited to, immune disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as AOSMC IL1B gb|X02532|HSIL1BR described herein under the heading “Immune Activity” and/or “Blood- VCAM gb|A30922|A30922 Related Disorders” (particularly including, but not limited to, immune disorders involving muscle tissues and the cardiovascular system (e.g. heart, lungs, circulatory system)). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving muscle tissue or the cardiovascular system). (AOSMC cells are human aortic smooth muscle cells). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as Daudi c-maf gb|AF055377|AF055377 described herein under the heading “Immune Activity” and/or “Blood- CD25 gb|X03137|HSIL2RG7 Related Disorders” (particularly including, but not limited to, immune CXCR3 gb|Z79783|HSCKRL2 disorders involving the B-cells). Highly preferred embodiments of the Granzyme B gb|J04071|HUMCSE invention include methods of preventing, detecting, diagnosing, treating ICAM gb|X06990|HSICAM1 and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving B-cells). (The Daudi cell line is a human B lymphoblast cell line available through the ATCC ™ as cell line number CCL-213). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as HEK293 CCR4 gb|AB023888|AB023888 described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). (The 293 cell line is a human embryonic kidney epithelial cell line available through the ATCC ™ as cell line number CRL-1573). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as HUVEC Rag2 gb|AY011962|AY011962 described herein under the heading “Immune Activity” and/or “Blood- VCAM gb|A30922|A30922 Related Disorders” (particularly including, but not limited to, immune disorders involving endothelial cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving endothelial cells). (HUVEC cells are human umbilical vein endothelial cells). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as Jurkat c-maf gb|AF055377|AF055377 described herein under the heading “Immune Activity” and/or “Blood- CD69 gb|Z22576|HSCD69GNA Related Disorders” (particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Jurkat cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number TIB-152). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as Liver VCAM gb|A30922|A30922 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving cells of the hepatic system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the hepatic system). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as NHDF HLA-c gb|AK027080|AK027080 described herein under the heading “Immune Activity” and/or “Blood- LTBR gb|M29474|HUMRAG1 Related Disorders” (particularly including, but not limited to, immune Rag1 disorders involving the skin). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the skin). (NHDF cells are normal human dermal fibroblasts). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as SK-N-MC CD40 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- neuroblastoma TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving the central nervous system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the central nervous system). (The SK-N-MC neuroblastoma cell line is a cell line derived from human brain tissue and is available through the ATCC ™ as cell line number HTB-10). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as T cell CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- CTLA4 gb|AF316875|AF316875 Related Disorders” (particularly including, but not limited to, immune Granzyme B gb|J04071|HUMCSE disorders involving T-cells). Highly preferred embodiments of the ICAM gb|X06990|HSICAM1 invention include methods of preventing, detecting, diagnosing, treating IFNg gb|X87308|HSRNAIG and/or ameliorating disorders of the immune system (particularly IL5 gb|X12705|HSBCDFIA including, but not limited to, immune disorders involving T-cells). LTBR gb|AK027080|AK027080 Rag2 gb|AY011962|AY011962 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as THP1 CCR3 gb|AB023887|AB023887 described herein under the heading “Immune Activity” and/or “Blood- CD30 gb|X04403|HS26KDAR Related Disorders” (particularly including, but not limited to, immune Il6 gb|AY011962|AY011962 disorders involving monocytes). Highly preferred embodiments of the Rag2 gb|A30922|A30922 invention include methods of preventing, detecting, diagnosing, treating VCAM and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The THP1 cell line is a human monocyte cell line available through the ATCC ™ as cell line number TIB-202). 105 HDPBQ71 Immune Highly preferred indications include immunological disorders such as U937 CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune VCAM gb|A30922|A30922 disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 184 HFCCQ50 Immune Highly preferred indications include immunological disorders such as TF-1 CD40 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- CD69 gb|Z22576|HSCD69GNA Related Disorders” (particularly including, but not limited to, immune disorders involving erythrocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving erythrocytes). (The TF-1 cell line is a human erythroblast cell line available through the ATCC ™ as cell line number CRL-2003). 184 HFCCQ50 Immune Highly preferred indications include immunological disorders such as U937 ICAM gb|X06990|HSICAM1 described herein under the heading “Immune Activity” and/or “Blood- IRF1 gb|X14454|HSIRF1 Related Disorders” (particularly including, but not limited to, immune LTBR gb|AK027080|AK027080 disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 186 HFCEW05 Immune Highly preferred indications include immunological disorders such as TF-1 CD40 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- IL1B gb|X02532|HSIL1BR Related Disorders” (particularly including, but not limited to, immune LTBR gb|AK027080|AK027080 disorders involving erythrocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving erythrocytes). (The TF-1 cell line is a human erythroblast cell line available through the ATCC ™ as cell line number CRL-2003). 204 HFVAB79 Immune Highly preferred indications include immunological disorders such as U937 CTLA4 gb|AF316875|AF316875 described herein under the heading “Immune Activity” and/or “Blood- ICAM gb|X06990|HSICAM1 Related Disorders” (particularly including, but not limited to, immune LTBR gb|AK027080|AK027080 disorders involving monocytes). Highly preferred embodiments of the TNF gb|AJ270944|HSA27094 invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as Adipocytes- ICAM gb|X06990|HSICAM1 described herein under the heading “Immune Activity” and/or “Blood- 3/12/01 Il6 gb|X04403|HS26KDAR Related Disorders” (particularly including, but not limited to, immune Rag1 gb|M29474|HUMRAG1 disorders involving adipocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving adipocytes). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as AOSMC CD30 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- CD40 gb|X02532|HSIL1BR Related Disorders” (particularly including, but not limited to, immune IL1B gb|X12705|HSBCDFIA disorders involving muscle tissues and the cardiovascular system (e.g. IL5 gb|AJ270944|HSA27094 heart, lungs, circulatory system)). Highly preferred embodiments of the TNF gb|A30922|A30922 invention include methods of preventing, detecting, diagnosing, treating VCAM and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving muscle tissue or the cardiovascular system). (AOSMC cells are human aortic smooth muscle cells). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as Caco-2 Rag1 gb|M29474|HUMRAG1 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving the cells of the gastrointestinal tract). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the gastrointestinal tract). (The Caco-2 cell line is a human colorectal adenocarcinoma cell line available through the ATCC ™ as cell line number HTB-37). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as Daudi ICAM gb|X06990|HSICAM1 described herein under the heading “Immune Activity” and/or “Blood- Rag1 gb|M29474|HUMRAG1 Related Disorders” (particularly including, but not limited to, immune VCAM gb|A30922|A30922 disorders involving the B-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving B-cells). (The Daudi cell line is a human B lymphoblast cell line available through the ATCC ™ as cell line number CCL-213). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as HEK293 c-maf gb|AF055377|AF055377 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). (The 293 cell line is a human embryonic kidney epithelial cell line available through the ATCC ™ as cell line number CRL-1573). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as HUVEC ICAM gb|X06990|HSICAM1 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving endothelial cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving endothelial cells). (HUVEC cells are human umbilical vein endothelial cells). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as Jurkat Rag2 gb|AY011962|AY011962 described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Jurkat cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number TIB-152). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as NHDF Rag1 gb|M29474|HUMRAG1 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving the skin). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the skin). (NHDF cells are normal human dermal fibroblasts). 247 HJACG02 Immune Highly preferred indications include immunological disorders such as U937 GATA1 gb|X17254|HSERYF1 described herein under the heading “Immune Activity” and/or “Blood- IL5 gb|X12705|HSBCDFIA Related Disorders”(particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 262 HKACD58 Immune Highly preferred indications include immunological disorders such as AOSMC VCAM gb|A30922|A30922 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving muscle tissues and the cardiovascular system (e.g. heart, lungs, circulatory system)). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving muscle tissue or the cardiovascular system). (AOSMC cells are human aortic smooth muscle cells). 262 HKACD58 Immune Highly preferred indications include immunological disorders such as Daudi CD40 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving the B-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving B-cells). (The Daudi cell line is a human B lymphoblast cell line available through the ATCC ™ as cell line number CCL-213). 262 HKACD58 Immune Highly preferred indications include immunological disorders such as HUVEC ICAM gb|X06990|HSICAM1 described herein under the heading “Immune Activity” and/or “Blood- Rag1 gb|M29474|HUMRAG1 Related Disorders” (particularly including, but not limited to, immune disorders involving endothelial cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving endothelial cells). (HUVEC cells are human umbilical vein endothelial cells). 262 HKACD58 Immune Highly preferred indications include immunological disorders such as Liver CD28 gb|AF222342|AF222342 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving cells of the hepatic system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the hepatic system). 262 HKACD58 Immune Highly preferred indications include immunological disorders such as NHDF CXCR3 gb|Z79783|HSCKRL2 described herein under the heading “Immune Activity” and/or “Blood- GATA1 gb|X17254|HSERYF1 Related Disorders” (particularly including, but not limited to, immune Il6 gb|X04403|HS26KDAR disorders involving the skin). Highly preferred embodiments of the VCAM gb|A30922|A30922 invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the skin). (NHDF cells are normal human dermal fibroblasts). 262 HKACD58 Immune Highly preferred indications include immunological disorders such as THP1 CIS3 gb|AB006967|AB006967 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The THP1 cell line is a human monocyte cell line available through the ATCC ™ as cell line number TIB-202). 262 HKACD58 Immune Highly preferred indications include immunological disorders such as U937 CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 277 HL2AC08 Immune Highly preferred indications include immunological disorders such as TF-1 CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- GATA1 gb|X17254|HSERYF1 Related Disorders” (particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving erythrocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving erythrocytes). (The TF-1 cell line is a human erythroblast cell line available through the ATCC ™ as cell line number CRL-2003). 383 HNHFO29 Immune Highly preferred indications include immunological disorders such as TF-1 CD40 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving erythrocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving erythrocytes). (The TF-1 cell line is a human erythroblast cell line available through the ATCC ™ as cell line number CRL-2003). 383 HNHFO29 Immune Highly preferred indications include immunological disorders such as U937 ICAM gb|X06990|HSICAM1 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as AOSMC CCR3 gb|AB023887|AB023887 described herein under the heading “Immune Activity” and/or “Blood- CCR4 gb|AB023888|AB023888 Related Disorders” (particularly including, but not limited to, immune CD25 gb|X03137|HSIL2RG7 disorders involving muscle tissues and the cardiovascular system (e.g. CD30 heart, lungs, circulatory system)). Highly preferred embodiments of the CD40 gb|AJ300189|HSA30018 invention include methods of preventing, detecting, diagnosing, treating CTLA4 gb|AF316875|AF316875 and/or ameliorating disorders of the immune system (particularly IL5 gb|X12705|HSBCDFIA including, but not limited to, immune disorders involving muscle tissue Rag1 gb|M29474|HUMRAG1 or the cardiovascular system). (AOSMC cells are human aortic smooth VCAM gb|A30922|A30922 muscle cells). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as Caco-2 c-maf gb|AF055377|AF055377 described herein under the heading “Immune Activity” and/or “Blood- GATA3 gb|X55037|HSGATA3 Related Disorders” (particularly including, but not limited to, immune ICAM gb|X06990|HSICAM1 disorders involving the cells of the gastrointestinal tract). Highly Rag1 gb|M29474|HUMRAG1 preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the gastrointestinal tract). (The Caco-2 cell line is a human colorectal adenocarcinoma cell line available through the ATCC ™ as cell line number HTB-37). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as Daudi TNF gb|AJ270944|HSA27094 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving the B-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving B-cells). (The Daudi cell line is a human B lymphoblast cell line available through the ATCC ™ as cell line number CCL-213). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as H9 CIS3 gb|AB006967|AB006967 described herein under the heading “Immune Activity” and/or “Blood- Rag1 gb|M29474|HUMRAG1 Related Disorders” (particularly including, but not limited to, immune disorders involving the T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The H9 cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number HTB-176). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as HEK293 CCR3 gb|AB023887|AB023887 described herein under the heading “Immune Activity” and/or “Blood- CCR4 gb|AB023888|AB023888 Related Disorders” (particularly including, but not limited to, immune CD25 gb|X03137|HSIL2RG7 disorders involving epithelial cells or the renal system). Highly CD30 preferred embodiments of the invention include methods of preventing, CD40 gb|AJ300189|HSA30018 detecting, diagnosing, treating and/or ameliorating disorders of the CTLA4 gb|AF316875|AF316875 immune system (particularly including, but not limited to, immune GATA3 gb|X55037|HSGATA3 disorders involving epithelial cells or the renal system). (The 293 cell Rag1 gb|M29474|HUMRAG1 line is a human embryonic kidney epithelial cell line available through TNF gb|AJ270944|HSA27094 the ATCC ™ as cell line number CRL-1573). VCAM gb|A30922|A30922 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as HUVEC CD40 gb|AJ300189|HSA30018 described herein under the heading “Immune Activity” and/or “Blood- ICAM gb|X06990|HSICAM1 Related Disorders” (particularly including, but not limited to, immune IL10 gb|AF055467|AF055467 disorders involving endothelial cells). Highly preferred embodiments of Rag1 gb|M29474|HUMRAG1 the invention include methods of preventing, detecting, diagnosing, Rag2 gb|AY011962|AY011962 treating and/or ameliorating disorders of the immune system TNF gb|AJ270944|HSA27094 (particularly including, but not limited to, immune disorders involving endothelial cells). (HUVEC cells are human umbilical vein endothelial cells). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as Jurkat CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- IL5 gb|X12705|HSBCDFIA Related Disorders” (particularly including, but not limited to, immune Rantes gb|AF043341|AF043341 disorders involving T-cells). Highly preferred embodiments of the TNF gb|AJ270944|HSA27094 invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Jurkat cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number TIB-152). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as Liver CD25 gb|X03137|HSIL2RG7 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving cells of the hepatic system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the hepatic system). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as Molt4 CD28 gb|AF222342|AF222342 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Molt-4 cell line is a human T-cell line available through the ATCC ™ as cell line number CRL-1582). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as NHDF CD28 gb|AF222342|AF222342 described herein under the heading “Immune Activity” and/or “Blood- CD40 gb|AJ300189|HSA30018 Related Disorders” (particularly including, but not limited to, immune Il6 gb|X04403|HS26KDAR disorders involving the skin). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the skin). (NHDF cells are normal human dermal fibroblasts). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as SK-N-MC c-maf gb|AF055377|AF055377 described herein under the heading “Immune Activity” and/or “Blood- neuroblastoma CIS3 gb|AB006967|AB006967 Related Disorders” (particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving the central nervous system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the central nervous system). (The SK-N-MC neuroblastoma cell line is a cell line derived from human brain tissue and is available through the ATCC ™ as cell line number HTB-10). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as SUPT TNF gb|AJ270944|HSA27094 described herein under the heading “Immune Activity” and/or “Blood- VCAM gb|A30922|A30922 Related Disorders” (particularly including, but not limited to, immune disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The SUPT cell line is a human T-cell line). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as THP1 CCR3 gb|AB023887|AB023887 described herein under the heading “Immune Activity” and/or “Blood- CD40 gb|AJ300189|HSA30018 Related Disorders” (particularly including, but not limited to, immune GATA3 gb|X55037|HSGATA3 disorders involving monocytes). Highly preferred embodiments of the ICAM gb|X06990|HSICAM1 invention include methods of preventing, detecting, diagnosing, treating IL5 gb|X12705|HSBCDFIA and/or ameliorating disorders of the immune system (particularly Rag2 gb|AY011962|AY011962 including, but not limited to, immune disorders involving monocytes). VCAM gb|A30922|A30922 (The THP1 cell line is a human monocyte cell line available through the ATCC ™ as cell line number TIB-202). 470 HSDSB09 Immune Highly preferred indications include immunological disorders such as U937 IL1B gb|X02532|HSIL1BR described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 562 HUKBT29 Immune Highly preferred indications include immunological disorders such as U937 CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as AOSMC CD30 described herein under the heading “Immune Activity” and/or “Blood- Il6 gb|X04403|HS26KDAR Related Disorders” (particularly including, but not limited to, immune disorders involving muscle tissues and the cardiovascular system (e.g. heart, lungs, circulatory system)). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving muscle tissue or the cardiovascular system). (AOSMC cells are human aortic smooth muscle cells). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as Caco-2 Rag1 gb|M29474|HUMRAG1 described herein under the heading “Immune Activity” and/or “Blood- Related Disorders” (particularly including, but not limited to, immune disorders involving the cells of the gastrointestinal tract). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the gastrointestinal tract). (The Caco-2 cell line is a human colorectal adenocarcinoma cell line available through the ATCC ™ as cell line number HTB-37). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as Daudi CIS3 gb|AB006967|AB006967 described herein under the heading “Immune Activity” and/or “Blood- CXCR3 gb|Z79783|HSCKRL2 Related Disorders” (particularly including, but not limited to, immune ICAM gb|X06990|HSICAM1 disorders involving the B-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving B-cells). (The Daudi cell line is a human B lymphoblast cell line available through the ATCC ™ as cell line number CCL-213). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as H9 IL5 gb|X12705|HSBCDFIA described herein under the heading “Immune Activity” and/or “Blood- VCAM gb|A30922|A30922 Related Disorders” (particularly including, but not limited to, immune VLA4 gb|X16983|HSINTAL4 disorders involving the T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The H9 cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number HTB-176). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as HEK293 Rag1 gb|M29474|HUMRAG1 described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving epithelial cells or the renal system). (The 293 cell line is a human embryonic kidney epithelial cell line available through the ATCC ™ as cell line number CRL-1573). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as HUVEC CCR7 gb|X84702|HSDNABLR2 described herein under the heading “Immune Activity” and/or “Blood- GATA3 gb|X55037|HSGATA3 Related Disorders” (particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving endothelial cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving endothelial cells). (HUVEC cells are human umbilical vein endothelial cells). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as Jurkat Rag1 gb|M29474|HUMRAG1 described herein under the heading “Immune Activity” and/or “Blood- Rag2 gb|AY011962|AY011962 Related Disorders” (particularly including, but not limited to, immune disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Jurkat cell line is a human T lymphocyte cell line available through the ATCC ™ as cell line number TIB-152). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as Liver CCR7 gb|X84702|HSDNABLR2 described herein under the heading “Immune Activity” and/or “Blood- ICAM gb|X06990|HSICAM1 Related Disorders” (particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving cells of the hepatic system). Highly preferred VCAM gb|A30922|A30922 embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving cells of the hepatic system). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as Molt4 CD25 gb|X03137|HSIL2RG7 described herein under the heading “Immune Activity” and/or “Blood- TNF gb|AJ270944|HSA27094 Related Disorders” (particularly including, but not limited to, immune VCAM gb|A30922|A30922 disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The Molt-4 cell line is a human T-cell line available through the ATCC ™ as cell line number CRL-1582). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as NHDF CCR7 gb|X84702|HSDNABLR2 described herein under the heading “Immune Activity” and/or “Blood- CD40 gb|AJ300189|HSA30018 Related Disorders” (particularly including, but not limited to, immune GATA3 gb|X55037|EISGATA3 disorders involving the skin). Highly preferred embodiments of the HLA-c invention include methods of preventing, detecting, diagnosing, treating TNF gb|AJ270944|HSA27094 and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the skin). (NHDF cells are normal human dermal fibroblasts). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as SK-N-MC CIS3 gb|AB006967|AB006967 described herein under the heading “Immune Activity” and/or “Blood- neuroblastoma LTBR gb|AK027080|AK027080 Related Disorders” (particularly including, but not limited to, immune Rag1 gb|M29474|HUMRAG1 disorders involving the central nervous system). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving the central nervous system). (The SK-N-MC neuroblastoma cell line is a cell line derived from human brain tissue and is available through the ATCC ™ as cell line number HTB-10). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as SUPT CCR4 gb|AB023888|AB023888 described herein under the heading “Immune Activity” and/or “Blood- Rag1 gb|M29474|HUMRAG1 Related Disorders” (particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving T-cells). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving T-cells). (The SUPT cell line is a human T-cell line). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as THP1 c-maf gb|AF055377|AF055377 described herein under the heading “Immune Activity” and/or “Blood- CCR7 gb|X84702|HSDNABLR2 Related Disorders” (particularly including, but not limited to, immune CXCR3 gb|Z79783|HSCKRL2 disorders involving monocytes). Highly preferred embodiments of the IL5 gb|X12705|HSBCDFIA invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The THP1 cell line is a human monocyte cell line available through the ATCC ™ as cell line number TIB-202). 584 HWHGZ51 Immune Highly preferred indications include immunological disorders such as U937 CD69 gb|Z22576|HSCD69GNA described herein under the heading “Immune Activity” and/or “Blood- ICAM gb|X06990|HSICAM1 Related Disorders” (particularly including, but not limited to, immune TNF gb|AJ270944|HSA27094 disorders involving monocytes). Highly preferred embodiments of the invention include methods of preventing, detecting, diagnosing, treating and/or ameliorating disorders of the immune system (particularly including, but not limited to, immune disorders involving monocytes). (The U937 cell line is a human monocyte cell line available through the ATCC ™ as cell line number CRL-1593.2).

Table 2 further characterizes certain encoded polypeptides of the invention, by providing the results of comparisons to protein and protein family databases. The first column provides a unique clone identifier, “Clone ID NO:”, corresponding to a cDNA clone disclosed in Table 1A and/or Table 1B. The second column provides the unique contig identifier, “Contig ID:” which allows correlation with the information in Table 1B. The third column provides the sequence identifier, “SEQ ID NO:”, for the contig polynucleotide sequences. The fourth column provides the analysis method by which the homology/identity disclosed in the Table was determined. The fifth column provides a description of the PFAM/NR hit identified by each analysis. Column six provides the accession number of the PFAM/NR hit disclosed in the fifth column. Column seven, score/percent identity, provides a quality score or the percent identity, of the hit disclosed in column five. Comparisons were made between polypeptides encoded by polynucleotides of the invention and a non-redundant protein database (herein referred to as “NR”), or a database of protein families (herein referred to as “PFAM”), as described below.

The NR database, which comprises the NBRF PIR database, the NCBI GenPept database, and the SIB SwissProt and TrEMBL databases, was made non-redundant using the computer program nrdb2 (Warren Gish, Washington University in Saint Louis). Each of the polynucleotides shown in Table 1B, column 3 (e.g., SEQ ID NO:X or the ‘Query’ sequence) was used to search against the NR database. The computer program BLASTX was used to compare a 6-frame translation of the Query sequence to the NR database (for information about the BLASTX algorithm please see Altshul et al., J. Mol. Biol. 215:403-410 (1990), and Gish and States, Nat. Genet. 3:266-272 (1993). A description of the sequence that is most similar to the Query sequence (the highest scoring ‘Subject’) is shown in column five of Table 2 and the database accession number for that sequence is provided in column six. The highest scoring ‘Subject’ is reported in Table 2 if (a) the estimated probability that the match occurred by chance alone is less than 1.0e-07, and (b) the match was not to a known repetitive element. BLASTX returns alignments of short polypeptide segments of the Query and Subject sequences which share a high degree of similarity; these segments are known as High-Scoring Segment Pairs or HSPs. Table 2 reports the degree of similarity between the Query and the Subject for each HSP as a percent identity in Column 7. The percent identity is determined by dividing the number of exact matches between the two aligned sequences in the HSP, dividing by the number of Query amino acids in the HSP and multiplying by 100. The polynucleotides of SEQ ID NO:X which encode the polypeptide sequence that generates an HSP are delineated by columns 8 and 9 of Table 2.

The PFAM database, PFAM version 2.1, (Sonnhammer, Nucl. Acids Res., 26:320-322, 1998)) consists of a series of multiple sequence alignments; one alignment for each protein family. Each multiple sequence alignment is converted into a probability model called a Hidden Markov Model, or HMM, that represents the position-specific variation among the sequences that make up the multiple sequence alignment (see, e.g., Durbin, et al., Biological sequence analysis: probabilistic models of proteins and nucleic acids, Cambridge University Press, 1998 for the theory of HMMs). The program HMMER version 1.8 (Sean Eddy, Washington University in Saint Louis) was used to compare the predicted protein sequence for each Query sequence (SEQ ID NO:Y in Table 1B) to each of the HMMs derived from PFAM version 2.1. A HMM derived from PFAM version 2.1 was said to be a significant match to a polypeptide of the invention if the score returned by HMMER 1.8 was greater than 0.8 times the HMMER 1.8 score obtained with the most distantly related known member of that protein family. The description of the PFAM family which shares a significant match with a polypeptide of the invention is listed in column 5 of Table 2, and the database accession number of the PFAM hit is provided in column 6. Column 7 provides the score returned by HMMER version 1.8 for the alignment. Columns 8 and 9 delineate the polynucleotides of SEQ ID NO:X which encode the polypeptide sequence which show a significant match to a PFAM protein family.

As mentioned, columns 8 and 9 in Table 2, “NT From” and “NT To”, delineate the polynucleotides of “SEQ ID NO:X” that encode a polypeptide having a significant match to the PFAM/NR database as disclosed in the fifth column. In one embodiment, the invention provides a protein comprising, or alternatively consisting of, a polypeptide encoded by the polynucleotides of SEQ ID NO:X delineated in columns 8 and 9 of Table 2. Also provided are polynucleotides encoding such proteins, and the complementary strand thereto.

The nucleotide sequence SEQ ID NO:X and the translated SEQ ID NO:Y are sufficiently accurate and otherwise suitable for a variety of uses well known in the art and described further below. For instance, the nucleotide sequences of SEQ ID NO:X are useful for designing nucleic acid hybridization probes that will detect nucleic acid sequences contained in SEQ ID NO:X or the cDNA contained in ATCC™ Deposit No:Z. These probes will also hybridize to nucleic acid molecules in biological samples, thereby enabling immediate applications in chromosome mapping, linkage analysis, tissue identification and/or typing, and a variety of forensic and diagnostic methods of the invention. Similarly, polypeptides identified from SEQ ID NO:Y may be used to generate antibodies which bind specifically to these polypeptides, or fragments thereof, and/or to the polypeptides encoded by the cDNA clones identified in, for example, Table 1A and/or 1B.

Nevertheless, DNA sequences generated by sequencing reactions can contain sequencing errors. The errors exist as misidentified nucleotides, or as insertions or deletions of nucleotides in the generated DNA sequence. The erroneously inserted or deleted nucleotides cause frame shifts in the reading frames of the predicted amino acid sequence. In these cases, the predicted amino acid sequence diverges from the actual amino acid sequence, even though the generated DNA sequence may be greater than 99.9% identical to the actual DNA sequence (for example, one base insertion or deletion in an open reading frame of over 1000 bases).

Accordingly, for those applications requiring precision in the nucleotide sequence or the amino acid sequence, the present invention provides not only the generated nucleotide sequence identified as SEQ ID NO:X, and a predicted translated amino acid sequence identified as SEQ ID NO:Y, but also a sample of plasmid DNA containing cDNA ATCC™ Deposit No:Z (e.g., as set forth in columns 2 and 3 of Table 1A and/or as set forth, for example, in Table 1B, 6, and 7). The nucleotide sequence of each deposited clone can readily be determined by sequencing the deposited clone in accordance with known methods. Further, techniques known in the art can be used to verify the nucleotide sequences of SEQ ID NO:X. The predicted amino acid sequence can then be verified from such deposits. Moreover, the amino acid sequence of the protein encoded by a particular clone can also be directly determined by peptide sequencing or by expressing the protein in a suitable host cell containing the deposited human cDNA, collecting the protein, and determining its sequence.

TABLE 2 PFam/NR Accession Score/Percent NT cDNA Clone ID Contig ID: SEQ ID NO: X Analysis Method PFam/NR Description Number Identity NT From To H6EAB28 589947 603 WUblastx.64 (Q9NXY7) CHONDROITIN 4-O-SULFOTRANSFERASE Q9NXY7 49% 205 396 (CHONDROITIN 4-O-SULFOTRANS 60% 1123 1206 100% 116 202 100% 1200 1352 38% 1118 1231 98% 413 1132 H6EDX46 637786 604 WUblastx.64 (Q9UHE9) ZSIG9 PROTEIN (TRANSMEMBRANE PROTEIN 4). Q9UHE9 100% 188 379 HACBD91 637482 17 WUblastx.64 NADH dehydrogenase (ubiquinone) (EC 1.6.5.3) chain NDUFB4 - pir|JE0383|JE0383 100% 211 357 human 95% 1306 1368 HACCI17 891114 18 HMMER 2.1.1 PFAM: PMP-22/EMP/MP20/Claudin family PF00822 142.7 470 1003 WUblastx.64 (AAH19290) Hypothetical 27.7 kDa protein (Fragment) AAH19290 100% 317 1114 HACCI17 731877 605 HMMER 2.1.1 PFAM: PMP-22/EMP/MP20/Claudin family PF00822 35.6 144 329 blastx.2 (AF000959) transmembrane protein [Homo sapiens] gb|AAC51364.1| 100% 311 619 93% 135 329 75% 535 786 HADAO89 570689 19 WUblastx.64 (Q9P147) PRO2822. Q9P147 73% 1106 885 HAGAI85 381942 21 WUblastx.64 (O15432) PROBABLE LOW-AFFINITY COPPER UPTAKE COP2_HUMAN 100% 91 234 PROTEIN 2 (HCT 96% 228 518 HAGAN21 1026956 23 WUblastx.64 (AAH07558) Unknown (protein for MGC: 15483). AAH07558 50% 797 708 46% 726 532 HAGBZ81 456414 24 WUblastx.64 (Q9H291) JUNCTATE. Q9H291 85% 183 329 77% 26 199 HAGDG59 534165 25 HMMER 2.1.1 PFAM: short chain dehydrogenase PF00106 182.2 232 795 WUblastx.64 (Q9UKU4) RETINAL SHORT-CHAIN Q9UKU4 96% 124 1023 DEHYDROGENASE/REDUCTASE RETSDR2. HAHDB16 635412 28 WUblastx.64 (Q9GMK2) HYPOTHETICAL 10.0 KDA PROTEIN. Q9GMK2 75% 641 522 69% 762 634 HAHDR32 635357 29 WUblastx.64 (Q9HBU9) POPEYE PROTEIN 2. Q9HBU9 92% 77 811 HAIBP89 727543 31 WUblastx.64 (Q96G79) Similar to RIKEN cDNA 2610030J16 gene. Q96G79 99% 290 1261 HAICP19 422672 32 WUblastx.64 (Q9H173) SIL1 PROTEIN PRECURSOR. Q9H173 100% 83 1465 HAJBR69 638516 35 WUblastx.64 (Q9JIG5) UBIQUITIN SPECIFIC PROTEASE (FRAGMENT). Q9JIG5 69% 677 48 HAJBZ75 618530 36 WUblastx.64 hypothetical protein DKFZp564D116.1 - human (fragment) pir|T08708|T08708 99% 25 1869 HAMFC93 904749 37 WUblastx.64 (Q9BGQ6) HYPOTHETICAL 30.3 KDA PROTEIN. Q9BGQ6 80% 1 666 HAMFC93 900586 611 WUblastx.64 (Q9BGQ6) HYPOTHETICAL 30.3 KDA PROTEIN. Q9BGQ6 94% 576 686 81% 4 573 HAPPW30 1352278 40 blastx.14 (AAH20263) Hypothetical 28.7 kDa protein. AAH20263 91% 59 850 HAPPW30 684272 613 WUblastx.64 (AAH20263) Hypothetical 28.7 kDa protein. AAH20263 100% 54 263 36% 982 1056 100% 266 844 HAPQT22 587601 41 WUblastx.64 (Q9H387) PRO2550. Q9H387 100% 462 439 68% 631 461 HASAV70 1300782 42 WUblastx.64 (Q9NY08) 19A PROTEIN. Q9NY08 82% 7 423 HASAV70 381953 614 WUblastx.64 (Q9NY08) 19A PROTEIN. Q9NY08 100% 4 432 HATAC53 1352276 44 blastx.14 (AAH19903) Hypothetical 29.4 kDa protein (Fragment) AAH19903 100% 64 699 100% 811 840 39% 1212 1280 HATAC53 667830 615 WUblastx.64 (AAH19903) Hypothetical 29.4 kDa protein (Fragment) AAH19903 98% 66 593 66% 516 665 HATBR65 635514 45 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 70% 750 610 68% 801 754 HATCP77 748244 47 WUblastx.64 (Q9Y691) MAXIK CHANNEL BETA 2 SUBUNIT (LARGE Q9Y691 100% 10 582 CONDUCTANCE CALCIUM-ACTI HATDF29 845965 48 WUblastx.64 (O95803) HEPARAN SULFATE N-DEACETYLASE/N- O95803 93% 143 1216 SULFOTRANSFERASE 3. HATDM46 974065 49 WUblastx.64 (AAH07609) Similar to hypothetical protein PRO1722. AAH07609 87% 2053 2030 82% 2274 2206 44% 2073 1945 68% 2210 2070 HBAFJ33 625916 51 WUblastx.64 (Q9GZR7) HYPOTHETICAL 96.3 KDA PROTEIN (ATP- Q9GZR7 96% 672 950 DEPENDENT RNA HELICASE) ( HBAFV19 843036 52 WUblastx.64 (Q9H068) HYPOTHETICAL 69.9 KDA PROTEIN. Q9H068 100% 3 779 HBCPB32 1045580 621 HMMER 2.1.1 PFAM: Sodium Bile acid symporter family PF01758 41.2 87 −230 WUblastx.64 (Q96EP9) Unknown (protein for IMAGE: 3502817) (Fragment). Q96EP9 48% 492 701 92% 2 589 HBHMA23 848016 56 WUblastx.64 (AAH08429) Similar to DNA segment, Chr 2, Massachus AAH08429 98% 643 1035 98% 71 649 HBIBW67 553678 57 WUblastx.64 (O60448) NEURONAL THREAD PROTEIN AD7C-NTP. O60448 71% 665 531 38% 617 384 67% 675 520 65% 896 837 40% 838 758 67% 1041 931 53% 699 430 71% 833 792 55% 894 838 31% 1077 871 46% 611 423 42% 835 743 31% 701 372 59% 530 465 38% 532 404 50% 1075 893 37% 523 404 46% 915 838 77% 1049 1023 61% 1042 785 HBIMB51 963208 58 WUblastx.64 (Q969E3) Urocortin III (Stresscopin). Q969E3 78% 98 535 HBIMB51 672711 623 WUblastx.64 (Q924A4) Urocortin III. Q924A4 61% 296 517 64% 93 302 HBINS58 1352386 59 blastx.14 (Q9D6W7) 2310047N01RIK PROTEIN. Q9D6W7 82% 255 578 64% 177 251 HBINS58 961712 624 WUblastx.64 (Q9D6W7) 2310047N01RIK PROTEIN. Q9D6W7 78% 191 589 HBINS58 892924 625 blastx.2 (AF106518) sialomucin CD164 [Homo sapiens] gb|AAC82473.1| 33% 241 576 HBJFU48 460392 60 WUblastx.64 (Q9P195) PRO1722. Q9P195 63% 716 660 73% 819 718 64% 667 533 HBJID05 1130660 61 WUblastx.64 probable ATP-binding component of ABC transporter PA4064 pir|H83138|H83138 74% 912 1604 [imported] - Pseudomonas aeruginosa (strain PAO1) HBJID05 544980 626 WUblastx.64 hypothetical protein PA4063 [imported] - Pseudomonas aeruginosa pir|G83138|G83138 61% 137 448 (strain PAO1) HBJIY92 778065 62 WUblastx.64 (Q9P529) Hypothetical 15.2 kDa protein. Q9P529 91% 2333 2434 86% 2325 2432 86% 2326 2433 100% 2348 2434 96% 2348 2434 86% 2344 2433 91% 2333 2434 HBJJU28 561723 63 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 61% 991 836 64% 1152 976 HBJLC01 638410 64 WUblastx.64 X-linked retinopathy protein (C-terminal, clone XEH.8c) - human pir|A46010|A46010 65% 829 707 (fragment) HBJLF01 732111 65 HMMER 2.1.1 PFAM: Transmembrane 4 family PF00335 131.8 223 891 WUblastx.64 (Q9D7W4) 2210021G21RIK PROTEIN. Q9D7W4 46% 133 894 HBJNC59 1125802 67 WUblastx.64 complement subcomponent C1q chain A precursor [validated] - human pir|S14350|C1HUQA 91% 66 800 HBJNC59 899397 627 HMMER 2.1.1 PFAM: Collagen triple helix repeat (20 copies) PF01391 30.1 144 245 WUblastx.64 (Q9H2L7) DC33. Q9H2L7 79% 77 907 HBJNC59 902207 628 HMMER 2.1.1 PFAM: C1q domain PF00386 250.2 409 786 blastx.2 (AF135157) complement C1q A chain precursor [Homo sapiens] gb|AAD32626.1| 91% 64 798 AF135157_1 HBOEG11 1300752 69 WUblastx.64 (O76076) CONNECTIVE TISSUE GROWTH FACTOR-LIKE O76076 75% 57 806 PROTEIN PRECURSOR (BA44 HBOEG11 1121709 629 HMMER 2.1.1 PFAM: Insulin-like growth factor binding proteins PF00219 45.4 128 340 WUblastx.64 (O76076) CONNECTIVE TISSUE GROWTH FACTOR-LIKE O76076 75% 53 802 PROTEIN PRECURSOR (BA44 HBOEG11 1049830 630 HMMER 2.1.1 PFAM: Insulin-like growth factor binding proteins PF00219 45.4 122 334 WUblastx.64 (O76076) CONNECTIVE TISSUE GROWTH FACTOR-LIKE O76076 100% 47 796 PROTEIN PRECURSOR (BA44 HBOEG69 793786 70 WUblastx.64 (Q9NS11) LIPOPOLYSACCHARIDE SPECIFIC RESPONSE-68 Q9NS11 71% 424 314 PROTEIN. 100% 345 196 HBXFL29 842802 71 WUblastx.64 (AAL36460) POB1. AAL36460 99% 4 1008 HCACU58 625923 72 WUblastx.64 (Q9NX85) CDNA FLJ20378 FIS, CLONE KAIA0536. Q9NX85 62% 497 820 HCACV51 1306706 73 WUblastx.64 (Q99LM9) UNKNOWN (PROTEIN FOR MGC: 8251). Q99LM9 85% 8 1009 HCACV51 598022 631 WUblastx.64 (Q96BN2) Similar to RIKEN cDNA 2900026B15 gene. Q96BN2 97% 13 312 100% 290 1015 HCDBW86 520435 74 WUblastx.64 (Q96P03) Transient receptor potential channel 4 zeta splice variant. Q96P03 100% 630 589 100% 463 371 HCE1Q89 520329 75 WUblastx.64 (Q9NX85) CDNA FLJ20378 FIS, CLONE KAIA0536. Q9NX85 86% 590 525 61% 645 592 65% 859 683 HCE2F54 634016 76 HMMER 2.1.1 PFAM: Histone-like transcription factor (CBF/NF-Y) and archaeal PF00808 19 868 1005 histone WUblastx.64 (AAH07642) Unknown (protein for IMAGE: 3534358) (Fra AAH07642 99% 298 1122 HCE3G69 728432 77 WUblastx.64 (Q9H0K7) HYPOTHETICAL 12.4 KDA PROTEIN (UNKNOWN) Q9H0K7 100% 1294 1647 (PROTEIN FOR MGC: 303 HCE3G69 494346 632 blastx.2 (AL136758) hypothetical protein [Homo sapiens] emb|CAB66692.1| 100% 1295 1648 HCEEA88 634967 78 WUblastx.64 (P57054) DOWN SYNDROME CRITICAL REGION PROTEIN 5 DSR5_HUMAN 100% 134 286 (DOWN SYND 99% 453 773 HCEFB69 748245 79 HMMER 2.1.1 PFAM: Mitochondrial carrier proteins PF00153 123.9 308 781 WUblastx.64 (Q9HC61) MITOCHONDRIAL UNCOUPLING PROTEIN 5 SHORT Q9HC61 96% 1093 1185 FORM WITH INSERTION 47% 1264 1320 98% 188 781 HCEFB80 1143407 80 WUblastx.64 (Q96FR3) Unknown (protein for MGC: 18083). Q96FR3 81% 1785 1979 HCEGR33 425212 81 WUblastx.64 (Q9H743) CDNA: FLJ21394 FIS, CLONE COL03536. Q9H743 51% 1002 1079 42% 1379 1492 58% 907 993 HCEMP62 684780 82 WUblastx.64 (AAL55739) Hypothetical 43.7 kDa protein. AAL55739 94% 484 897 94% 88 459 40% 1 198 94% 870 926 HCEWE17 941941 84 WUblastx.64 (Q9H310) RH TYPE B GLYCOPROTEIN. Q9H310 84% 9 293 92% 444 566 HCEWE17 893535 635 WUblastx.64 (Q9H310) RH TYPE B GLYCOPROTEIN. Q9H310 80% 467 544 75% 695 730 83% 3 482 HCEWE20 543370 85 WUblastx.64 (Q9P1J1) PRO1546. Q9P1J1 76% 501 551 79% 601 717 HCFOM18 553582 89 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 60% 621 490 HCHNF25 658672 637 WUblastx.64 (AAH00499) Jumping translocation breakpoint. AAH00499 91% 180 620 HCMST14 562010 92 WUblastx.64 (Q96DD7) Hypothetical 24.9 kDa protein (Fragment). Q96DD7 100% 10 99 HCMTB45 862367 93 WUblastx.64 (Q9UI48) PRO0663 (FRAGMENT). Q9UI48 67% 748 656 61% 952 914 HCMTB45 562034 638 WUblastx.64 (Q9UI48) PRO0663 (FRAGMENT). Q9UI48 67% 741 649 61% 945 907 HCOOS80 1134974 95 WUblastx.64 (Q60838) SEGMENT POLARITY PROTEIN DISHEVELLED DVL2_MOUSE 47% 8 307 HOMOLOG DVL-2 42% 440 637 100% 636 677 HCOOS80 1045182 639 blastx.2 similar to Dvl-1 product encoded by GenBank Accession Number 1 gb|AAC52827.1| 100% 21 128 100% 588 683 39% 80 163 36% 8 94 45% 444 509 HCUCK44 720291 98 WUblastx.64 hypothetical protein DKFZp564J157.1 - human (fragment) pir|T34520|T34520 97% 21 524 HCUEO60 499242 99 WUblastx.64 (Q96MM0) CDNA FLJ32172 fis, clone PLACE6000555. Q96MM0 79% 1043 972 72% 1222 1028 HCUHK65 651313 101 WUblastx.64 (Q9H3W5) HYPOTHETICAL 79.4 KDA PROTEIN. Q9H3W5 100% 11 316 HCUHK65 880178 642 HMMER 2.1.1 PFAM: Leucine Rich Repeat PF00560 92.1 1190 1261 WUblastx.64 (Q9H3W5) HYPOTHETICAL 79.4 KDA PROTEIN. Q9H3W5 100% 770 2893 HCWEB58 1352416 103 blastx.14 (Q92WW6) Putative sensor histidine kinase protein. Q92WW6 36% 355 720 51% 946 1167 55% 853 933 41% 264 335 37% 757 828 HCWEB58 1115089 643 HMMER 2.1.1 PFAM: Domain found in bacterial signal proteins PF00672 40.4 442 651 WUblastx.64 sensor histidine kinase [imported] - Caulobacter crescentus pir|A87396|A87396 36% 379 915 HCWEB58 889268 644 HMMER 2.1.1 PFAM: Domain found in bacterial signal proteins PF00672 41.6 350 559 blastx sensor-like protein [Coxiella burnetii] gb|AAA81939.1| 39% 419 829 HCWGU37 1042325 104 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 86% 2588 2523 77% 2459 2409 77% 2521 2441 73% 2373 2329 70% 2730 2587 HCWKC15 553621 105 WUblastx.64 (Q9NX85) CDNA FLJ20378 FIS, CLONE KAIA0536. Q9NX85 77% 538 419 56% 710 663 63% 708 532 HDHEB60 499233 107 WUblastx.64 (Q9Y5Y5) PEROXISOMAL BIOGENESIS FACTOR 16. Q9Y5Y5 81% 277 1284 HDHIA94 765171 108 HMMER 2.1.1 PFAM: Sodium/calcium exchanger protein PF01699 121.4 178 615 WUblastx.64 (Q9HC58) SODIUM/CALCIUM EXCHANGER NCKX3. Q9HC58 98% 10 657 HDHIA94 637576 646 HMMER 2.1.1 PFAM: Sodium/calcium exchanger protein PF01699 22.9 187 273 blastx.2 (AF025664) Na—Ca + K exchanger [Bos taurus] gb|AAB88884.1| 42% 224 637 37% 16 273 HDHMA45 902513 109 WUblastx.64 (Q9H3S3) TRANSMEMBRANE PROTEASE, SERINE 5 (EC 3.4.21.—) TMS5_HUMAN 91% 175 1428 (SP HDHMA45 812764 647 HMMER 2.1.1 PFAM: Trypsin PF00089 296.3 723 1415 WUblastx.64 (Q9H3S3) TRANSMEMBRANE PROTEASE, SERINE 5 (EC 3.4.21.—) TMS5_HUMAN 99% 180 1442 (SP HDHMA72 547772 110 WUblastx.64 (AAH17663) Hypothetical 55.1 kDa protein. AAH17663 28% 3700 3891 95% 761 1168 50% 1019 1231 99% 2 592 HDLAC10 692299 111 WUblastx.64 (Q9UBJ4) TRANSPOSASE-LIKE PROTEIN. Q9UBJ4 99% 29 1378 HDPBA28 1062783 113 WUblastx.64 (Q9UKY2) ADIPOCYTE-DERIVED LEUCINE AMINOPEPTIDASE. Q9UKY2 94% 259 3081 HDPBA28 866429 648 HMMER 2.1.1 PFAM: Peptidase family M1 PF01433 613.6 228 1391 WUblastx.64 (Q9UKY2) ADIPOCYTE-DERIVED LEUCINE AMINOPEPTIDASE. Q9UKY2 99% 69 2891 HDPBQ02 745403 649 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 45% 1667 1548 60% 2188 2021 51% 1741 1643 61% 2022 1846 HDPBQ71 1160316 115 WUblastx.64 (Q9BRE2) HYPOTHETICAL 68.4 KDA PROTEIN (FRAGMENT). Q9BRE2 100% 90 1928 HDPBQ71 727200 650 WUblastx.64 (Q9BRE2) HYPOTHETICAL 68.4 KDA PROTEIN (FRAGMENT). Q9BRE2 99% 21 1859 HDPBQ71 886067 651 WUblastx.64 (Q9H2V9) CDA08. Q9H2V9 100% 1532 1999 65% 169 264 44% 182 322 21% 1456 1551 93% 186 1541 HDPCY37 837699 117 HMMER 2.1.1 PFAM: Glycosyl hydrolase family 47 PF01532 627.5 199 1521 WUblastx.64 (Q9H886) CDNA FLJ13869 FIS, CLONE THYRO1001287, WEAKLY Q9H886 92% 76 1809 SIMILAR TO MAN HDPCY37 604114 652 HMMER 2.1.1 PFAM: Glycosyl hydrolase family 47 PF01532 324 199 834 HDPFF39 588697 118 WUblastx.64 (O96005) CLEFT LIP AND PALATE TRANSMEMBRANE O96005 100% 3 29 PROTEIN 1. 100% 97 762 HDPGK25 704067 119 WUblastx.64 (Q9H6I8) CDNA: FLJ22233 FIS, CLONE HRC02016. Q9H6I8 98% 3 701 HDPGP94 823355 120 WUblastx.64 (Q14288) HYPOTHETICAL PROTEIN (FRAGMENT). Q14288 50% 689 216 39% 1093 890 55% 909 700 27% 1804 1652 27% 1767 1090 29% 2282 2082 HDPJF37 704487 122 WUblastx.64 (Q9BSQ8) UNKNOWN (PROTEIN FOR IMAGE: 3510191) Q9BSQ8 94% 105 650 (FRAGMENT). 36% 158 718 93% 19 153 HDPJM30 879325 123 WUblastx.64 (O94759) LONG TRANSIENT RECEPTOR POTENTIAL CHANNEL TRL2_HUMAN 99% 17 1633 2 (LTRPC HDPJM30 603517 653 WUblastx.64 (O94759) LONG TRANSIENT RECEPTOR POTENTIAL CHANNEL TRL2_HUMAN 89% 416 1312 2 (LTRPC 96% 378 530 98% 1 378 HDPNC61 637585 124 WUblastx.64 (AAG23169) HC6. AAG23169 52% 654 827 64% 37 78 HDPND46 637586 125 WUblastx.64 (Q9BR26) DJ257E24.3 (NOVEL PROTEIN) (FRAGMENT). Q9BR26 98% 12 1466 HDPOE32 897276 126 WUblastx.64 (Q9BW48) MY047 PROTEIN. Q9BW48 98% 64 345 HDPOH06 683371 127 HMMER 2.1.1 PFAM: Uncharacterized membrane protein family PF01554 90.8 255 596 WUblastx.64 (Q96FL8) Hypothetical 61.9 kDa protein. Q96FL8 99% 18 977 HDPOZ56 815653 654 HMMER 2.1.1 PFAM: Flavin containing amine oxidase PF01593 431.1 307 1614 WUblastx.64 (Q96RQ9) Interleukin-4 induced gene-1 protein. Q96RQ9 99% 103 1800 HDPOZ56 743479 655 HMMER 2.1.1 PFAM: Flavin containing amine oxidase PF01593 185.2 200 949 HDPPA04 904765 129 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 19.1 373 582 WUblastx.64 (Q9BQ51) BUTYROPHILIN PRECURSOR B7-DC (PD-1-LIGAND 2 Q9BQ51 92% 271 1089 PROTEIN). HDPPH47 630030 130 WUblastx.64 (Q9NUN5) CDNA FLJ11240 FIS, CLONE PLACE1008568. Q9NUN5 99% 116 1735 HDPSB18 1043263 131 WUblastx.64 (Q9H5R3) CDNA: FLJ23147 FIS, CLONE LNG09295. Q9H5R3 70% 3363 3163 HDPSP01 689129 661 WUblastx.64 (Q9BR97) UNKNOWN (PROTEIN FOR MGC: 10763). Q9BR97 90% 227 1114 98% 1078 1668 100% 1664 1744 HDPTD15 692917 135 WUblastx.64 (Q9BU29) UNKNOWN (PROTEIN FOR IMAGE: 3954899) Q9BU29 97% 937 833 (FRAGMENT). HDPTK41 744824 136 WUblastx.64 (BAB11849) MOP-2. BAB11849 97% 1013 1126 99% 72 1025 HDPUG50 684120 137 WUblastx.64 (O60860) GLUCOSYLTRANSFERASE (FRAGMENT). O60860 99% 4 1599 HDPUH26 866433 138 WUblastx.64 (Q9NXE5) CDNA FLJ20296 FIS, CLONE HEP05890. Q9NXE5 100% 735 1736 HDPUW68 812737 139 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 38.9 844 1005 WUblastx.64 (Q9Y286) QA79 MEMBRANE PROTEIN, ALLELIC VARIANT Q9Y286 95% 70 1440 AIRM-1B PRECURSOR. HDPVH60 796865 140 WUblastx.64 (BAB55096) CDNA FLJ14508 fis, clone NT2RM1000421, w BAB55096 95% 229 294 54% 229 294 40% 214 294 25% 1137 1544 27% 981 1622 29% 987 1478 29% 1215 1487 31% 1389 1607 38% 288 473 94% 1580 2077 99% 288 1610 23% 291 1544 25% 1607 2029 27% 1604 2017 30% 1658 2032 24% 660 1469 45% 229 294 34% 154 282 28% 813 1598 HDPWN93 992925 141 WUblastx.64 (Q9H747) CDNA: FLJ21347 FIS, CLONE COL02724. Q9H747 99% 201 2450 HDQHD03 1309175 142 WUblastx.64 (O60859) NEUROPATHY TARGET ESTERASE. O60859 68% 382 1149 HDQHD03 834692 665 HMMER 2.1.1 PFAM: Cyclic nucleotide-binding domain PF00027 44.3 709 870 WUblastx.64 (O60859) NEUROPATHY TARGET ESTERASE. O60859 63% 259 1134 HDTBP04 1307742 143 WUblastx.64 (Q9D5J3) 4930432K09RIK PROTEIN. Q9D5J3 46% 70 369 HDTBP04 543618 666 WUblastx.64 (Q9D5J3) 4930432K09RIK PROTEIN. Q9D5J3 38% 65 718 HDTFE17 1043391 146 WUblastx.64 (Q9UJU8) JM24 PROTEIN (FRAGMENT). Q9UJU8 100% 14 118 84% 955 1089 100% 343 705 HDTFE17 892317 671 HMMER 2.1.1 PFAM: Transmembrane amino acid transporter protein PF01490 86.4 116 481 HDTIT10 839264 148 HMMER 2.1.1 PFAM: Phosphatidylethanolamine-binding protein PF01161 50.9 463 621 WUblastx.64 (Q96DV4) Similar to RIKEN cDNA 4733401F03 gene. Q96DV4 100% 816 911 91% 352 858 HDTIT10 834697 672 blastx.2 (AF117272) O-crystallin [Octopus dofleini] gb|AAD29640.1| 31% 337 765 AF117272_1 HDTMK50 1011485 149 WUblastx.64 (Q9P195) PRO1722. Q9P195 53% 1119 1030 72% 1186 1121 76% 1331 1176 HE2DY70 722217 150 WUblastx.64 (Q9BS33) SIMILAR TO HYPOTHETICAL PROTEIN FLJ11218. Q9BS33 100% 9 167 HE2FV03 396139 152 WUblastx.64 (CAD13327) BA382H24.3 (multiple PDZ domain protein) CAD13327 73% 281 805 HE2NV57 740750 153 WUblastx.64 (Q9UGV6) BK445C9.3 (HIGH-MOBILITY GROUP (NONHISTONE Q9UGV6 31% 321 866 CHROMOSOMAL) PROT 66% 71 106 HE2PD49 638617 154 WUblastx.64 (Q9BSR6) SIMILAR TO RIKEN CDNA 2410018G23 GENE. Q9BSR6 100% 403 849 HE8MH91 589450 157 WUblastx.64 (Q9H8Z4) CDNA FLJ13121 FIS, CLONE NT2RP3002687. Q9H8Z4 98% 9 410 HE8QV67 1050076 158 WUblastx.64 (BAB55430) CDNA FLJ14978 fis, clone VESEN1000122. BAB55430 100% 321 425 31% 487 600 100% 1 201 84% 1403 1684 74% 577 729 86% 800 1108 HE8UB86 834913 159 WUblastx.64 (Q9D390) 6330503C03RIK PROTEIN. Q9D390 44% 103 189 61% 432 707 HE9BK23 675382 160 HMMER 2.1.1 PFAM: Fibrinogen beta and gamma chains, C-terminal globular domain PF00147 77.2 762 959 WUblastx.64 (Q9Y5C1) ANGIOPOIETIN 5. Q9Y5C1 100% 958 1419 98% 39 959 HE9CO69 596829 161 WUblastx.64 (O95772) H_NH1021A08.1 PROTEIN (UNKNOWN) (PROTEIN FOR O95772 100% 25 291 MGC: 14607) (SIM HE9DG49 1299935 163 WUblastx.64 (Q9NYL4) FK506 BINDING PROTEIN PRECURSOR. Q9NYL4 93% 127 618 HE9DG49 658678 676 HMMER 2.1.1 PFAM: FKBP-type peptidyl-prolyl cis-trans isomerases PF00254 91 211 492 WUblastx.64 (Q9NYL4) FK506 BINDING PROTEIN PRECURSOR. Q9NYL4 100% 70 672 HE9DG49 382000 677 HMMER 2.1.1 PFAM: FKBP-type peptidyl-prolyl cis-trans isomerases PF00254 91 −71 −352 HE9OW20 838598 678 WUblastx.64 (CAC41349) Alpha2-glucosyltransferase. CAC41349 99% 136 1059 HE9OW20 834400 679 blastx.2 potassium channel regulator 1 [Rattus norvegicus] gb|AAC34249.1| 81% 449 1051 90% 129 497 HE9RM63 886167 165 WUblastx.64 (Q9NV86) CDNA FLJ10873 FIS, CLONE NT2RP4001730, WEAKLY Q9NV86 40% 1995 2087 SIMILAR TO UDP 100% 82 1113 HEAAR07 561524 166 WUblastx.64 probable transposase - human transposon MER37 pir|S72481|S72481 57% 691 858 89% 1020 1076 75% 210 332 78% 833 1015 33% 784 864 55% 332 703 HEBCM63 484643 169 WUblastx.64 (Q9BYH1) SEZ6L. Q9BYH1 91% 12 449 HEBEJ18 701802 170 WUblastx.64 (AAH00573) HSPC163 protein. AAH00573 100% 51 467 HEEAG23 684254 171 HMMER 2.1.1 PFAM: emp24/gp25L/p24 family PF01105 36.2 63 185 WUblastx.64 (Q9CZL0) 2400003B06RIK PROTEIN. Q9CZL0 59% 3 185 80% 406 780 HEEAJ02 633657 172 WUblastx.64 (Q9BW86) PHOSPHATIDYLETHANOLAMINE N- Q9BW86 99% 54 761 METHYLTRANSFERASE. HEEAQ11 777843 173 HMMER 2.1.1 PFAM: Cystatin domain PF00031 39.7 360 638 WUblastx.64 (Q9H4G1) BA218C14.1 (NOVEL CYSTATIN FAMILY MEMBER). Q9H4G1 87% 213 653 HEGAN94 885637 174 WUblastx.64 colipase precursor, pancreatic - dog pir|A46717|A46717 36% 148 393 HEGAN94 769649 680 HMMER 2.1.1 PFAM: Colipase PF01114 24 229 405 WUblastx.64 colipase precursor, pancreatic - dog pir|A46717|A46717 36% 229 474 HELGK31 681138 176 HMMER 2.1.1 PFAM: DHHC zinc finger domain PF01529 95.1 659 820 WUblastx.64 (Q9NPG8) CDNA FLJ10479 FIS, CLONE NT2RP2000120 (DC1) Q9NPG8 99% 209 1240 (HYPOTHETICAL 39 HELGK31 340352 682 HMMER 2.1.1 PFAM: DHHC zinc finger domain PF01529 95.1 −82 −243 blastx.2 CDNA FLJ10479 FIS, CLONE NT2RP2000120 (DC1). sp|Q9NPG8|Q9NPG8 100% 498 1274 98% 242 496 36% 36 128 HELHD85 847372 177 WUblastx.64 (Q9N083) UNNAMED PORTEIN PRODUCT. Q9N083 52% 1715 1653 53% 1648 1559 67% 1881 1705 HELHL48 696945 178 HMMER 2.1.1 PFAM: DHHC zinc finger domain PF01529 124.3 797 991 WUblastx.64 hypothetical protein DKFZp761E1347.1 - human (fragment) pir|T47144|T47144 100% 359 1501 HELHL48 610025 683 HMMER 2.1.1 PFAM: DHHC zinc finger domain PF01529 124.3 199 393 WUblastx.64 hypothetical protein DKFZp761E1347.1 - human (fragment) pir|T47144|T47144 100% 470 586 99% 585 905 100% 10 471 HEMAM41 741647 179 WUblastx.64 (AAH21428) Hypothetical 20.0 kDa protein. AAH21428 67% 175 744 HEPAA46 596830 180 WUblastx.64 (Q96PH6) ESC42. Q96PH6 100% 18 386 HEQCC55 884824 685 WUblastx.64 (Q9NP84) TYPE I TRANSMENMBRANE PROTEIN PRECURSOR Q9NP84 64% 62 397 (TYPE I TRANSMEMBRAN HESAJ10 526013 185 WUblastx.64 (AAK95397) Selenoprotein SelM. AAK95397 96% 566 841 100% 477 545 72% 550 582 HETAB45 609827 186 WUblastx.64 (Q9NXH2) CDNA FLJ20254 FIS, CLONE COLF6926. Q9NXH2 98% 646 795 99% 3 647 HETEU28 1018676 188 WUblastx.64 (Q96CP0) Similar to old astrocyte specifically induced substance. Q96CP0 94% 7 714 HETLM70 1177512 189 WUblastx.64 (Q9H766) CDNA: FLJ21240 FIS, CLONE COL01132. Q9H766 40% 3 989 HETLM70 1046327 688 blastx.2 B0416.1 gene product [Caenorhabditis elegans] gb|AAB36841.2| 30% 231 977 HFABG18 847073 190 WUblastx.64 (Q9QZE9) TM6P1. Q9QZE9 95% 53 253 88% 237 797 HFABH95 566712 191 WUblastx.64 (Q9QZH5) PUTATIVE PHOSPHATE/PHOSPHOENOLPYRUVATE Q9QZH5 88% 513 944 TRANSLOCATOR. 65% 9 77 HFAMB72 490697 192 WUblastx.64 (Q9Y6F6) JAW1-RELATED PROTEIN MRVI1A LONG ISOFORM. Q9Y6F6 94% 672 722 69% 1 669 HFCCQ50 579993 194 HMMER2.1.1 PFAM: Galactosyltransferase PF01762 130.8 365 1042 WUblastx.64 (Q9C0J1) BETA-1,3-N-ACETYLGLUCOSAMINYLTRANSFERASE Q9C0J1 95% 35 1102 BGN-T4. HFCDK17 381980 195 WUblastx.64 (Q9H2Q4) HT027. Q9H2Q4 80% 388 540 97% 488 910 HFFAL36 560639 198 WUblastx.64 (O75525) T-STAR. O75525 100% 568 657 HFGAD82 513669 199 WUblastx.64 membrane glycoprotein M6 - mouse pir|I78556|I78556 92% 249 410 HFIIN69 1011487 200 WUblastx.64 (Q9UI86) PRO0113. Q9UI86 55% 1448 1281 HFIIZ70 1043350 201 WUblastx.64 (AAK95397) Selenoprotein SelM. AAK95397 100% 990 1145 92% 842 919 96% 102 182 91% 423 458 HFKET18 889515 202 WUblastx.64 (Q9HAD8) CDNA FLJ11786 FIS, CLONE HEMBA1006036. Q9HAD8 63% 1384 1485 54% 1230 1397 42% 1444 1533 66% 1390 1434 50% 1471 1533 HFPAO71 629193 207 WUblastx.64 (O60448) NEURONAL THREAD PROTEIN AD7C-NTP. O60448 69% 1201 998 60% 2034 1846 60% 2009 1791 53% 1223 1026 40% 1223 1134 32% 1786 1601 44% 993 940 59% 1059 934 63% 2022 1789 35% 2040 1708 32% 1935 1690 36% 1126 941 44% 1141 932 42% 1144 953 39% 1275 1150 56% 1834 1697 64% 1220 996 HFPCX09 1309793 208 WUblastx.64 (O95970) LEUCINE-RICH GLIOMA-INACTIVATED PROTEIN O95970 96% 161 1831 PRECURSOR. HFPCX09 835390 694 HMMER 2.1.1 PFAM: Leucine rich repeat C-terminal domain PF01463 46.3 741 890 WUblastx.64 (O95970) LEUCINE-RICH GLIOMA-INACTIVATED PROTEIN O95970 99% 225 1895 PRECURSOR. HFPCX09 598723 695 blastx.2 (AF055636) leucine-rich glioma-inactivated protein precursor [Homo gb|AAC99316.1| 94% 169 1830 sapiens] 86% 161 298 HFPCX36 526635 209 WUblastx.64 (Q96NR6) CDNA FLJ30278 fis, clone BRACE2002755. Q96NR6 56% 680 775 66% 450 680 HFPCX64 1309796 210 WUblastx.64 (Q9H743) CDNA: FLJ21394 FIS, CLONE COL03536. Q9H743 54% 893 783 56% 1066 872 HFPCX64 877637 696 WUblastx.64 (Q9H743) CDNA: FLJ21394 FIS, CLONE COL03536. Q9H743 54% 891 781 56% 1064 870 HFPCX64 514187 698 blastx.2 (AF000996) ubiquitous TPR motif, Y isoform [Homo sapiens] gb|AAC51843.1| 56% 1125 859 HFTBM50 545012 212 WUblastx.64 (Q9H8P0) CDNA FLJ13352 FIS, CLONE OVARC1002165, Q9H8P0 100% 23 229 WEAKLY SIMILAR TO 3-O 91% 198 524 HFTDL56 695976 213 HMMER2.1.1 PFAM: Neurotransmitter-gated ion-channel PF00065 769.9 168 1574 WUblastx.64 (P04760) ACETYLCHOLINE RECEPTOR PROTEIN, GAMMA ACHG_MOUSE 99% 93 1649 CHAIN PRECUR HFVAB79 1300736 214 WUblastx.64 (Q9BX93) GROUP XIII SECRETED PHOSPHOLIPASE A2. Q9BX93 100% 133 693 HFVAB79 565076 699 WUblastx.64 (Q9BX93) GROUP XIII SECRETED PHOSPHOLIPASE A2. Q9BX93 100% 139 720 HFXGT26 745381 218 WUblastx.64 (O95662) POT. ORF VI (FRAGMENT). O95662 57% 162 689 HFXJU68 570855 700 WUblastx.64 (Q9CW46) 1300006N24RIK PROTEIN (FRAGMENT). Q9CW46 51% 9 599 HFXKJ03 505207 222 WUblastx.64 (O62658) LINE-1 ELEMENT ORF2. O62658 34% 492 292 36% 920 525 HFXKY27 634161 223 WUblastx.64 (Q9H743) CDNA: FLJ21394 FIS, CLONE COL03536. Q9H743 87% 763 716 63% 821 765 46% 936 814 HGBFO79 422794 224 WUblastx.64 (AAH06833) Similar to DKFZP586F1524 protein. AAH06833 78% 72 140 96% 134 1147 HGBIB74 837220 226 WUblastx.64 hypothetical protein ZK858.6 - Caenorhabditis elegans pir|T28058|T28058 50% 1387 1494 51% 2 439 65% 482 730 62% 723 1403 HGBIB74 838602 701 blastx.2 Similar to S. cerevisiae EMP70 protein precursor (S25110) [Homo dbj|BAA13385.1| 87% 736 1257 sapiens] 100% 79 477 78% 1251 1505 88% 537 749 HGBIB74 899864 702 blastx.2 Similar to S. cerevisiae EMP70 protein precursor (S25110) [Homo dbj|BAA13385.1| 87% 12 950 sapiens] HHAAF20 838603 228 WUblastx.64 (Q9NXG9) CDNA FLJ20259 FIS, CLONE COLF7443 Q9NXG9 87% 540 728 (HYPOTHETICAL 47.5 KDA PRO 81% 245 580 HHEAA08 638231 229 WUblastx.64 (Q9BVD9) UNKNOWN (PROTEIN FOR MGC: 5149). Q9BVD9 61% 1923 1870 73% 2123 1923 HHEBB10 604124 230 WUblastx.64 (Q9NQR1) PR/SET DOMAIN CONTAINING PROTEIN 07. Q9NQR1 95% 2 136 HHEMM74 941955 233 WUblastx.64 (Q96QU0) Calcium-promoted Ras inactivator. Q96QU0 87% 1741 2046 HHEMM74 906815 704 blastx unknown [Homo sapiens] gb|AAC50940.1| 61% 830 738 53% 713 636 71% 731 711 HHENK42 493724 234 WUblastx.64 (AAK55521) PRO0764. AAK55521 54% 644 441 HHENP27 799532 235 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 26.5 120 353 WUblastx.64 (Q96BF3) Hypothetical 30.7 kDa protein. Q96BF3 99% 12 857 HHEPM33 877639 238 WUblastx.64 (Q96BH1) Ring finger protein 25. Q96BH1 97% 10 1230 100% 1185 1373 HHEPU04 838217 240 WUblastx.64 (Q9BQB6) UNKNOWN (PROTEIN FOR MGC: 11276) (PROTEIN Q9BQB6 100% 259 747 FOR IMAGE: 3455200). HHEPU04 897457 707 blastx.2 (BC000828) Unknown (protein for IMAGE: 3455200) [Homo sapiens] gb|AAH00828.1| 80% 267 755 AAH00828 HHEPU04 535730 708 WUblastx.64 (Q9BQB6) UNKNOWN (PROTEIN FOR MGC: 11276) (PROTEIN Q9BQB6 72% 326 424 FOR IMAGE: 3455200). 83% 217 339 100% 45 218 HHFEC49 905849 241 WUblastx.64 (Q9D1N2) 1110002J19RIK PROTEIN. Q9D1N2 56% 180 500 HHFGR93 865581 242 WUblastx.64 (Q96AP7) Hypothetical 41.2 kDa protein. Q96AP7 100% 132 1301 HHFGR93 691402 709 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 36.3 628 807 WUblastx.64 (Q96AP7) Hypothetical 41.2 kDa protein. Q96AP7 98% 819 1298 99% 130 828 HHFHR32 411470 244 WUblastx.64 (Q99LX9) SIMILAR TO SINGLE-STRANDED-DNA-BINDING Q99LX9 100% 1 762 PROTEIN. HHFOJ29 1127491 245 WUblastx.64 (Q9H7P4) FLJ00024 PROTEIN (FRAGMENT). Q9H7P4 99% 592 65 HHGCM76 662329 246 WUblastx.64 (Q96FV2) Unknown (protein for IMAGE: 3945715) (Fragment). Q96FV2 94% 7 114 98% 378 536 HHGCM76 383547 712 WUblastx.64 (Q96FV2) Unknown (protein for IMAGE: 3945715) (Fragment). Q96FV2 94% 7 114 98% 378 536 HHGDW43 554613 248 WUblastx.64 (Q9P1J1) PRO1546. Q9P1J1 59% 707 787 52% 774 887 HHPGO40 1299927 250 WUblastx.64 (Q9HBW1) Brain tumor associated protein NAG14. Q9HBW1 74% 191 976 30% 338 928 HHPGO40 753270 713 HMMER 2.1.1 PFAM: Leucine Rich Repeat PF00560 122 542 613 WUblastx.64 (Q9HBW1) Brain tumor associated protein NAG14. Q9HBW1 74% 191 967 30% 338 928 HHPGO40 560969 714 HMMER 2.1.1 PFAM: Leucine Rich Repeat PF00560 77 548 619 HHSGW69 1031514 253 WUblastx.64 (O95325) PROTEASOME SUBUNIT P58. O95325 100% 730 780 94% 529 582 HHTLF25 461438 254 WUblastx.64 (Q9UMT3) KILLER ACTIVATING RECEPTOR ASSOCIATED Q9UMT3 91% 142 474 PROTEIN, ISOFORM B. HJABX32 487807 255 WUblastx.64 (O70277) RING FINGER PROTEIN. O70277 98% 463 612 45% 198 449 35% 75 440 37% 339 458 50% 25 72 35% 463 588 34% 463 582 38% 523 600 31% 108 476 84% 3 458 HJACA79 562729 256 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 63% 642 577 73% 880 644 HJACG02 1307789 257 WUblastx.64 (Q9HD89) CYSTEINE-RICH SECRETED PROTEIN (C/EBP- Q9HD89 100% 111 389 EPSILON REGULATED MYEL HJACG02 509948 717 WUblastx.64 (Q9HD89) CYSTEINE-RICH SECRETED PROTEIN (C/EBP- Q9HD89 100% 47 370 EPSILON REGULATED MYEL HJACG30 895505 258 WUblastx.64 (Q9UM21) UDP-GLCNAC:A-1,3-D-MANNOSIDE B-1,4-N- Q9UM21 96% 291 389 ACETYLGLUCOSAMINYLTRANS HJBAV55 823510 259 WUblastx.64 (Q9H360) PRO1331. Q9H360 100% 1202 1393 HJBCU04 877643 260 WUblastx.64 (Q9Y3P8) SIT PROTEIN PRECURSOR. Q9Y3P8 100% 36 623 HJPAD75 651337 265 WUblastx.64 (Q9H5F8) CDNA: FLJ23476 FIS, CLONE HSI14935. Q9H5F8 98% 8 232 HKAAE44 564406 266 WUblastx.64 (Q969S6) Unknown (protein for MGC: 15961) (protein for Q969S6 99% 113 520 MGC: 14327). HKAAH36 836040 722 WUblastx.64 (AAH08036) Kallikrein 5. AAH08036 90% 184 348 100% 399 1061 HKAAH36 838068 723 HMMER 2.1.1 PFAM: Trypsin PF00089 270.2 452 1108 WUblastx.64 (AAH08036) Kallikrein 5. AAH08036 100% 254 1132 HKAAH36 815661 724 HMMER 2.1.1 PFAM: Trypsin PF00089 270.2 327 983 WUblastx.64 (AAH08036) Kallikrein 5. AAH08036 92% 129 1007 HKAAK02 589945 268 HMMER 2.1.1 PFAM: Galactosyltransferase PF01762 56.1 457 660 WUblastx.64 (CAC82374) Beta 1,6-GlcNAc-transferase. CAC82374 92% 97 681 HKABZ65 862030 270 WUblastx.64 (Q96LB9) Peptidoglycan recognition protein-I-alpha precursor. Q96LB9 90% 77 802 39% 137 541 HKABZ65 665424 726 WUblastx.64 (Q96LB9) Peptidoglycan recognition protein-I-alpha precursor. Q96LB9 99% 69 794 45% 129 533 HKACB56 554616 271 HMMER 2.1.1 PFAM: Kazal-type serine protease inhibitor domain PF00050 76.3 114 266 WUblastx.64 (P01001) ACROSIN INHIBITORS IIA AND IIB (BUSI-II). IAC2_BOVIN 82% 96 266 HKACD58 552465 727 WUblastx.64 (Q96BH2) Hypothetical 34.4 kDa protein. Q96BH2 86% 795 1208 87% 122 724 HKACM93 1352383 273 blastx.14 aqualysin (EC 3.4.21.—) I precursor - Thermus aquaticus pir|A35742|A35742 40% 884 1039 41% 1097 1276 30% 1274 1468 50% 746 823 34% 548 670 53% 425 469 58% 2201 2236 HKADQ91 604123 274 WUblastx.64 (Q9NWC5) HYPOTHETICAL 31.7 KDA PROTEIN. Q9NWC5 100% 229 1053 HKAEG43 889521 275 WUblastx.64 (Q9NRD1) F-BOX PROTEIN FBG2. Q9NRD1 100% 204 1082 HKAEL80 570865 276 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 72% 935 1000 75% 1002 1073 61% 763 957 HKAEV06 638238 733 WUblastx.64 (Q9NVA4) CDNA FLJ10846 FIS, CLONE NT2RP4001373. Q9NVA4 96% 367 459 100% 197 367 96% 480 1541 HKAFK41 545018 278 WUblastx.64 (BAB55101) CDNA FLJ14515 fis, clone NT2RM1000800, w BAB55101 91% 18 371 60% 130 537 HKDBF34 833065 279 WUblastx.64 (Q9HBJ8) KIDNEY-SPECIFIC MEMBRANE PROTEIN NX-17. Q9HBJ8 88% 69 734 HKDBF34 587268 734 WUblastx.64 (Q9HBJ8) KIDNEY-SPECIFIC MEMBRANE PROTEIN NX-17. Q9HBJ8 100% 18 257 96% 239 682 HKGAT94 762811 280 WUblastx.64 (Q9H919) CDNA FLJ13078 FIS, CLONE NT2RP3002002. Q9H919 73% 307 239 80% 128 84 63% 228 121 HKGAT94 460631 735 blastx.2 pval [Plasmodium vivax] emb|CAA63219.1| 41% 456 154 62% 148 116 HKISB57 625956 282 WUblastx.64 (AAL36150) Smoothelin-B3. AAL36150 28% 262 582 100% 201 1013 98% 1107 1256 27% 271 480 26% 532 966 44% 954 1052 HKIYP40 580845 284 WUblastx.64 (Q9H7S6) CDNA FLJ14310 FIS, CLONE PLACE3000271. Q9H7S6 47% 468 295 61% 1098 1036 77% 1215 1189 HKMLP68 1037919 286 WUblastx.64 (AAH17691) Hypothetical 61.8 kDa protein. AAH17691 42% 8 586 HL2AC08 610018 287 HMMER 2.1.1 PFAM: Thioredoxin PF00085 82.8 145 444 WUblastx.64 hypothetical protein DKFZp564E1962.1 - human (fragment) pir|T12471|T12471 80% 46 903 HLCND09 1172046 289 HMMER 2.1.1 PFAM: PAP2 superfamily PF01569 20.3 170 352 WUblastx.64 (Q9H929) CDNA FLJ13055 FIS, CLONE NT2RP3001538, WEAKLY Q9H929 88% 107 421 SIMILAR TO HYP HLCND09 1035153 739 HMMER 2.1.1 PFAM: PAP2 superfamily PF01569 20.4 62 244 blastx.2 (AK000307) unnamed protein product [Homo sapiens] dbj|BAA91072.1| 50% 2 325 HLDBX13 815665 290 WUblastx.64 (Q9H387) PRO2550. Q9H387 76% 1764 1681 60% 1815 1756 HLDOW79 847396 292 WUblastx.64 (Q90YM5) Organic solute transporter alpha. Q90YM5 47% 10 672 HLDQC46 847397 293 WUblastx.64 (Q9BXJ8) TRANSMEMBRANE PROTEIN INDUCED BY TUMOR Q9BXJ8 100% 28 423 NECROSIS FACTOR ALPHA HLDQR62 753742 294 WUblastx.64 (Q9NQW2) PROGRESSIVE ANKYLOSIS-LIKE PROTEIN. Q9NQW2 100% 41 382 99% 376 1002 HLDQU79 740755 295 WUblastx.64 (O75477) KE04P. O75477 100% 105 1142 HLDRM43 846330 296 WUblastx.64 (Q96NZ9) Proline-rich acidic protein. Q96NZ9 92% 24 476 HLDRM43 638939 740 WUblastx.64 (Q96NZ9) Proline-rich acidic protein. Q96NZ9 100% 164 616 HLDRP33 647430 297 WUblastx.64 (Q9H743) CDNA: FLJ21394 FIS, CLONE COL03536. Q9H743 38% 340 278 64% 599 489 HLHFP03 460467 298 WUblastx.64 (Q9WVC2) LY-6/NEUROTOXIN HOMOLOG (ADULT MALE Q9WVC2 81% 224 571 HIPPOCAMPUS CDNA, RIKEN HLICQ90 791828 301 WUblastx.64 (Q96N65) CDNA FLJ31349 fis, clone MESAN2000092, moderately Q96N65 95% 571 636 similar to 93% 59 616 HLJBJ61 1019012 302 WUblastx.64 deoxyhypusine synthase (EC 1.1.1.249) [validated] - human pir|S68692|S68692 93% 805 1047 HLMCA59 519349 304 WUblastx.64 (Q9NX17) CDNA FLJ20489 FIS, CLONE KAT08285. Q9NX17 75% 783 649 HLQBE09 520375 305 WUblastx.64 second peroxisomal thioesterase - human pir|JC7367|JC7367 56% 8 559 HLQDH79 588446 306 WUblastx.64 (Q9HBZ6) HT005 PROTEIN. Q9HBZ6 100% 404 556 HLQDR48 1307726 307 WUblastx.64 (Q9NQZ1) HEPATOCELLULAR CARCINOMA ASSOCIATED Q9NQZ1 86% 296 406 PROTEIN TD26. HLQDR48 619979 745 WUblastx.64 (Q9NQZ1) HEPATOCELLULAR CARCINOMA ASSOCIATED Q9NQZ1 83% 289 399 PROTEIN TD26. HLQEM64 897823 746 WUblastx.64 (Q9NVB5) CDNA FLJ10829 FIS, CLONE NT2RP4001138. Q9NVB5 99% 3 437 HLTAU74 853614 309 WUblastx.64 (AAH21123) Hypothetical 113.9 kDa protein (Fragment AAH21123 93% 6 704 37% 6 803 HLTCO33 778074 310 WUblastx.64 (Q96MM0) CDNA FLJ32172 fis, clone PLACE6000555. Q96MM0 72% 982 917 62% 1179 973 HLTHG37 787530 314 WUblastx.64 (AAH01258) N-acetylglucosamine-phosphate mutase. AAH01258 100% 960 1070 93% 2 955 HLWAA17 629552 315 WUblastx.64 (Q9NY26) IRT1 PROTEIN (SIMILAR TO ZINC/IRON REGULATED Q9NY26 99% 85 960 TRANSPORTER-LIK HLWAD77 653513 316 WUblastx.64 (Q9GZP9) F-LAN-1 (HYPOTHETICAL TRANSMEMBRANE Q9GZP9 99% 29 745 PROTEIN SBBI53). HLWAE11 783071 317 HMMER 2.1.1 PFAM: C1q domain PF00386 44.4 403 789 WUblastx.64 (Q9BXI9) COMPLEMENT-C1Q TUMOR NECROSIS FACTOR- Q9BXI9 99% 28 861 RELATED PROTEIN. HLWAO22 587270 318 WUblastx.64 (Q9NRG9) GL003 (ADRACALIN) (AAAS PROTEIN) (UNKNOWN) Q9NRG9 78% 449 1147 (PROTEIN FOR MGC: 28% 139 420 97% 1003 1263 100% 14 40 83% 19 495 30% 396 596 41% 503 664 28% 100 408 26% 470 859 58% 333 503 HLWAY54 658702 319 WUblastx.64 (Q9BY87) PROACROSIN BINDING PROTEIN SP32 PRECURSOR. Q9BY87 78% 38 1006 100% 1448 1663 100% 1251 1448 23% 1445 1594 37% 1260 1331 80% 1006 1326 HLWBY76 797609 321 WUblastx.64 (AAH06651) Similar to hypothetical protein FLJ23153 AAH06651 76% 6 1127 HLYAN59 553507 748 WUblastx.64 (Q9P529) Hypothetical 15.2 kDa protein. Q9P529 93% 631 720 96% 638 721 90% 631 720 96% 638 721 96% 638 721 92% 638 721 76% 620 721 89% 638 721 96% 638 721 100% 638 721 HLYAZ61 423998 749 HMMER 2.1.1 PFAM: 7 transmembrane receptor (rhodopsin family) PF00001 71.8 254 −309 WUblastx.64 (O14626) PROBABLE G PROTEIN-COUPLED RECEPTOR H963. H963_HUMAN 98% 1 846 HMADS41 596831 328 WUblastx.64 (AAH07725) Ceroid-lipofuscinosis, neuronal 8 (epile AAH07725 92% 186 449 100% 427 1041 HMADU73 467053 750 WUblastx.64 (Q9EPE8) LOW-DENSITY LIPOPROTEIN RECEPTOR-RELATED Q9EPE8 78% 115 294 PROTEIN 9. HMAMI15 1352406 330 blastx.14 (Q96QY4) BA134O15.1 (similar to citrate lyase) (Fragment). Q96QY4 99% 85 1023 HMAMI15 1049263 751 WUblastx.64 (Q96QY4) BA134015.1 (similar to citrate lyase) (Fragment). Q96QY4 79% 372 920 100% 84 440 HMDAE65 520338 331 WUblastx.64 (Q9NLE3) PROBABLE (HHV-6) U1102, VARIANT A DNA, Q9NLE3 79% 335 249 COMPLETE VIRION GENOM 70% 342 250 63% 333 235 64% 330 235 67% 333 250 HMECK83 636035 335 WUblastx.64 (O62658) LINE-1 ELEMENT ORF2. O62658 32% 668 483 50% 65 6 49% 483 100 HMEED18 560775 336 WUblastx.64 (Q9H651) CDNA: FLJ22604 FIS, CLONE HSI04630 (BBP-LIKE Q9H651 93% 34 696 PROTEIN 2). HMEET96 566720 337 WUblastx.64 (Q9CR48) 2610318G18RIK PROTEIN. Q9CR48 86% 121 915 HMIAL37 603201 338 HMMER 2.1.1 PFAM: PDZ domain (Also known as DHR or GLGF). PF00595 57.7 127 327 WUblastx.64 (Q9Y6N9) ANTIGEN NY-CO-38. Q9Y6N9 100% 315 1100 100% 76 315 38% 109 318 27% 870 1061 35% 765 998 62% 1111 1242 63% 1067 1132 HMIAP86 726831 339 HMMER 2.1.1 PFAM: Mitochondrial carrier proteins PF00153 262 329 1180 WUblastx.64 (AAG29582) Mitochondrial uncoupling protein 5 long AAG29582 97% 182 1183 HMMAH60 562776 341 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 52% 675 538 53% 820 665 HMQDT36 1309723 343 WUblastx.64 (Q9D1Q6) 1110001E24RIK PROTEIN. Q9D1Q6 89% 157 1350 HMQDT36 424085 753 HMMER 2.1.1 PFAM: Thioredoxin PF00085 59.8 76 −265 WUblastx.64 (Q9D1Q6) 1110001E24RIK PROTEIN. Q9D1Q6 93% 192 1409 HMSBX80 597448 344 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 61% 1721 1413 HMSGB14 570833 346 WUblastx.64 (Q9BGV8) HYPOTHETICAL 10.0 KDA PROTEIN. Q9BGV8 73% 403 615 HMSGU01 1049069 347 WUblastx.64 (Q9H8U5) CDNA FLJ13219 FIS, CLONE NT2RP4001849, WEAKLY Q9H8U5 59% 418 840 SIMILAR TO SH3 HMSGU01 1158803 754 WUblastx.64 (Q9H8U5) CDNA FLJ13219 FIS, CLONE NT2RP4001849, WEAKLY Q9H8U5 45% 217 657 SIMILAR TO SH3 82% 221 838 HMSGU01 853368 755 WUblastx.64 (Q9H8U5) CDNA FLJ13219 FIS, CLONE NT2RP4001849, WEAKLY Q9H8U5 100% 215 838 SIMILAR TO SH3 HMSHS36 1127691 349 WUblastx.64 (O95662) POT. ORF VI (FRAGMENT). O95662 83% 781 350 HMSHS36 1028961 756 blastx.2 pot. ORF VI [Homo sapiens] emb|CAA26920.1| 61% 539 378 77% 609 490 HMSKC04 799540 352 WUblastx.64 (Q9H743) CDNA: FLJ21394 FIS, CLONE COL03536. Q9H743 66% 1341 1225 60% 1414 1346 56% 1244 1053 HMTAD67 588447 353 WUblastx.64 (Q9N083) UNNAMED PORTEIN PRODUCT. Q9N083 64% 1171 947 HMUAP70 872208 354 WUblastx.64 (Q9EQH8) NEDD4 WW DOMAIN-BINDING PROTEIN 5 Q9EQH8 89% 69 845 (FRAGMENT). HMUAP70 778820 758 WUblastx.64 (Q9BT67) UNKNOWN (PROTEIN FOR MGC: 10924). Q9BT67 100% 183 221 72% 229 402 100% 338 844 HMUAP70 381964 761 blastx.2 (AF220209) Nedd4 WW domain-binding protein 5 [Mus musculus] gb|AAG44248.1| 86% 1 720 HMWEB02 638159 356 WUblastx.64 (Q96MX0) CDNA FLJ31762 fis, clone NT2RI2007754, weakly similar Q96MX0 100% 10 207 to INT 34% 187 300 97% 333 479 HMWFO02 542061 762 WUblastx.64 (Q9P1C6) PRO2738. Q9P1C6 61% 647 549 44% 473 345 HMWFY10 825421 358 WUblastx.64 (Q9NYF4) PUTATIVE ZINC FINGER PROTEIN. Q9NYF4 98% 14 226 HMWFY10 490495 763 WUblastx.64 (Q9NYF4) PUTATIVE ZINC FINGER PROTEIN. Q9NYF4 98% 14 226 HMWGY65 1308287 359 WUblastx.64 (Q9D624) 1200003C23RIK PROTEIN. Q9D624 55% 42 1442 HMWGY65 794987 764 WUblastx.64 (AAH19452) Hypothetical 49.0 kDa protein. AAH19452 58% 542 1438 65% 42 596 HNEEB45 1036397 361 WUblastx.64 hypothetical protein 3 - human pir|E41925|E41925 78% 861 929 39% 523 717 44% 566 862 HNFFC43 753337 362 WUblastx.64 (Q96BY8) Hypothetical 55.2 kDa protein. Q96BY8 97% 319 453 66% 428 769 87% 651 839 99% 903 1517 HNFJF07 577013 364 WUblastx.64 (AAL55831) Hypothetical 14.1 kDa protein. AAL55831 65% 585 457 HNFJH45 410107 365 WUblastx.64 (Q9H7Z0) CDNA FLJ14058 FIS, CLONE HEMBB1000554. Q9H7Z0 48% 277 11 HNGAK47 561488 366 WUblastx.64 (Q96EF8) Unknown (protein for MGC: 21495). Q96EF8 33% 12 206 31% 12 206 20% 492 617 34% 492 557 25% 486 569 39% 190 2 29% 537 487 HNGEG08 494246 372 WUblastx.64 (Q9P0U4) CPG BINDING PROTEIN. Q9P0U4 64% 270 67 HNGEP09 499076 374 WUblastx.64 (AAK55521) PRO0764. AAK55521 57% 965 861 53% 1021 977 50% 867 715 HNGIJ31 519120 377 WUblastx.64 (Q9N083) UNNAMED PORTEIN PRODUCT. Q9N083 73% 566 610 54% 615 725 66% 454 561 HNGJE50 561568 379 WUblastx.64 (Q9HBS7) HYPOTHETICAL 14.2 KDA PROTEIN. Q9HBS7 64% 1028 945 62% 919 734 HNGJP69 604891 381 WUblastx.64 (Q9H743) CDNA: FLJ21394 FIS, CLONE COL03536. Q9H743 53% 973 857 71% 860 693 HNGOM56 836064 384 WUblastx.64 (Q96MM0) CDNA FLJ32172 fis, clone PLACE6000555. Q96MM0 38% 577 744 58% 714 953 HNHFO29 463568 393 WUblastx.64 (Q9NX85) CDNA FLJ20378 FIS, CLONE KAIA0536. Q9NX85 69% 522 695 HNHOD46 843488 395 WUblastx.64 (O60448) NEURONAL THREAD PROTEIN AD7C-NTP. O60448 76% 334 552 56% 646 921 56% 645 713 52% 844 894 73% 331 498 59% 353 625 50% 828 917 70% 721 792 48% 781 915 50% 558 791 35% 401 595 31% 283 552 50% 379 462 61% 486 839 HNTBL27 545534 397 WUblastx.64 (Q96AA3) Putative endoplasmic reticulum multispan transmembrane Q96AA3 98% 243 500 prote 33% 13 168 40% 646 711 96% 13 261 HNTCE26 1160395 398 HMMER 2.1.1 PFAM: 7 transmembrane receptor (rhodopsin family) PF00001 137.5 282 1037 WUblastx.64 (Q9H1Y3) DJ317G22.2 (ENCEPHALOPSIN) (PANOPSIN). Q9H1Y3 100% 111 1316 HNTCE26 853373 774 HMMER 2.1.1 PFAM: 7 transmembrane receptor (rhodopsin family) PF00001 23.2 63 218 WUblastx.64 (Q9H1Y3) DJ317G22.2 (ENCEPHALOPSIN) (PANOPSIN). Q9H1Y3 95% 370 495 100% 12 377 HOACB38 520201 401 WUblastx.64 (Q9H387) PRO2550. Q9H387 71% 420 295 77% 589 419 HODDN65 520348 403 WUblastx.64 (Q9N083) UNNAMED PORTEIN PRODUCT. Q9N083 74% 743 663 67% 660 493 HODDN92 422913 404 WUblastx.64 (Q9H1S5) BA110H4.2 (SIMILAR TO MEMBRANE PROTEIN). Q9H1S5 100% 1119 1021 HODDO08 790333 405 WUblastx.64 (AAL55740) Hypothetical 11.9 kDa protein. AAL55740 100% 725 1042 HODDW40 579256 406 WUblastx.64 (Q9NX85) CDNA FLJ20378 FIS, CLONE KAIA0536. Q9NX85 59% 677 495 HODGE68 834907 408 WUblastx.64 retrovirus-related hypothetical protein II - human 1 pir|S23650|S23650 36% 370 278 54% 276 1 HOEBK34 768325 409 HMMER 2.1.1 PFAM: von Willebrand factor type C domain PF00093 54.1 455 619 WUblastx.64 (O94769) EXTRACELLULAR MATRIX PROTEIN. O94769 96% 149 643 HOEBK34 509951 778 WUblastx.64 (O94769) EXTRACELLULAR MATRIX PROTEIN. O94769 96% 68 325 93% 316 561 HOEBZ89 828177 410 WUblastx.64 hypothetical protein C05G5.5 - Caenorhabditis elegans pir|T18967|T18967 31% 133 1008 HOEDB32 634994 411 WUblastx.64 (Q9Y2Y6) TADA1 PROTEIN (DKFZP564K1964 PROTEIN). Q9Y2Y6 95% 104 781 HOEDE28 1036480 412 WUblastx.64 (AAG23764) PP3686. AAG23764 99% 933 1535 HOEDH84 748236 413 WUblastx.64 (Q960D8) SD05564p. Q960D8 39% 7 1449 HOFMQ33 1184465 414 WUblastx.64 (O15232) MATRILIN-3 PRECURSOR. MTN3_HUMAN 85% 205 1500 HOFMQ33 919896 780 HMMER 2.1.1 PFAM: von Willebrand factor type A domain PF00092 189.8 288 815 WUblastx.64 (O15232) MATRILIN-3 PRECURSOR. MTN3_HUMAN 85% 204 1499 HOFMQ33 906694 781 HMMER 2.1.1 PFAM: von Willebrand factor type A domain PF00092 162.2 318 737 HOFMT75 911180 415 HMMER 2.1.1 PFAM: Eukaryotic aspartyl protease PF00026 619 290 1303 WUblastx.64 cathepsin D (EC 3.4.23.5) precursor [validated] - human pir|A25771|KHHUD 81% 83 1303 HOFMT75 892291 786 HMMER 2.1.1 PFAM: Eukaryotic aspartyl protease PF00026 496.2 336 1232 WUblastx.64 cathepsin D (EC 3.4.23.5) precursor [validated] - human pir|A25771|KHHUD 96% 192 1232 HOFND85 847424 417 HMMER 2.1.1 PFAM: Cadherin domain PF00028 256 905 1180 WUblastx.64 (AAK51617) Protocadherin-beta7. AAK51617 83% 167 2047 30% 425 1858 HOFOC33 1186156 419 WUblastx.64 clusterin precursor - dog pir|A40018|A40018 69% 1022 1414 81% 115 1086 HOFOC33 967554 788 HMMER 2.1.1 PFAM: Clusterin PF01093 236.4 81 395 WUblastx.64 clusterin precursor - dog pir|A40018|A40018 65% 120 449 HOFOC33 878690 789 HMMER 2.1.1 PFAM: Clusterin PF01093 236.6 81 395 HOFOC33 905734 790 HMMER 2.1.1 PFAM: Clusterin PF01093 301.2 76 432 blastx.2 glycoprotein 80 [Canis familiaris] gb|AAA30846.1| 81% 440 1087 69% 1023 1415 81% 115 432 HOFOC33 806819 793 HMMER 2.1.1 PFAM: 60s Acidic ribosomal protein PF00428 74.6 −422 −733 WUblastx.64 acidic ribosomal protein P0, cytosolic [validated] - human pir|A27125|R5HUP0 52% 5 55 87% 42 812 HOGCK20 745445 420 WUblastx.64 (Q969N2) Phosphatidyl inositol glycan class T precursor (Hypothetical Q969N2 99% 378 1622 97% 57 389 HOGCK63 895880 421 WUblastx.64 (Q9Y386) CGI-78 PROTEIN. Q9Y386 69% 1214 1252 88% 1161 1214 92% 514 1161 HOGCK63 902295 795 WUblastx.64 (Q96BI3) Hypothetical 29.0 kDa protein. Q96BI3 100% 813 872 96% 22 477 HOGCS52 919898 422 WUblastx.64 (Q9NY68) CTL2 PROTEIN. Q9NY68 90% 79 1383 HOHBB49 833080 423 WUblastx.64 (Q96MM0) CDNA FLJ32172 fis, clone PLACE6000555. Q96MM0 57% 2582 2292 HOHBC68 603968 424 WUblastx.64 (AAH20256) Hypothetical 110.4 kDa protein. AAH20256 94% 348 707 97% 676 1785 HOHCH55 827481 427 WUblastx.64 (O95965) TEN INTEGRIN EGF-LIKE REPEAT DOMAINS O95965 84% 221 1702 PROTEIN PRECURSOR. HOHCH55 815682 798 blastx.2 (AF072752) ten integrin EGF-like repeat domains protein precursor gb|AAD17666.1| 99% 230 1621 [Homo sapiens] 42% 326 1426 35% 416 1576 100% 1623 1712 HOSDJ25 854234 428 WUblastx.64 (Q9D8Y9) 1810018L05RIK PROTEIN. Q9D8Y9 85% 468 593 86% 143 544 HOSEG51 545809 429 WUblastx.64 (Q9NUT5) CDNA FLJ11152 FIS, CLONE PLACE1006901 Q9NUT5 51% 2 82 (FRAGMENT). 100% 46 537 HOSEQ49 588824 430 WUblastx.64 (Q9UP47) TNF-INDUCED PROTEIN GG2-1. Q9UP47 99% 120 683 HOSFD58 614040 431 HMMER 2.1.1 PFAM: ATP-sulfurylase PF01747 697.3 −335 −1321 WUblastx.64 3′-phosphoadenosine-5′-phosphosulfate synthetase - human pir|JW0087|JW0087 100% 56 1927 HOUCQ17 429229 432 HMMER 2.1.1 PFAM: Reprolysin family propeptide PF01562 76.2 216 −20 WUblastx.64 (P97857) ADAM-TS 1 PRECURSOR (EC 3.4.24.—) (A DISINTEGRIN A ATS1_MOUSE 81% 508 3408 HOUDK26 565393 433 WUblastx.64 (Q9NUX1) CDNA FLJ11082 FIS, CLONE PLACE1005206. Q9NUX1 94% 4 585 HOUGG12 775824 802 WUblastx.64 (Q9H2K4) DM4E3. Q9H2K4 94% 619 1290 41% 688 1437 28% 536 703 27% 619 1194 33% 649 1266 28% 778 1269 100% 413 616 22% 649 1212 HPASA81 900548 803 HMMER 2.1.1 PFAM: CUB domain PF00431 146.9 452 778 WUblastx.64 (O35360) UTERUS-OVARY SPECIFIC PUTATIVE O35360 70% 8 928 TRANSMEMBRANE PROTEIN. 75% 918 1814 HPASA81 801923 804 blastx.2 (AF022147) uterus-ovary specific putative transmembrane protein gb|AAB71895.1| 69% 299 1924 [Rattus norvegicus] 60% 106 333 33% 641 934 40% 1009 1119 HPBCU51 411080 437 WUblastx.64 (Q9BWJ9) SIMILAR TO NEUROBLASTOMA (NERVE TISSUE) Q9BWJ9 96% 56 154 PROTEIN. HPFCL43 535710 440 WUblastx.64 (AAH07349) Adrenal gland protein AD-004. AAH07349 97% 57 257 HPFDG48 542227 441 WUblastx.64 (Q9Y6E5) HSPC024-ISO. Q9Y6E5 90% 564 623 88% 313 387 HPIAQ68 833082 442 WUblastx.64 (Q95LL4) Hypothetical 13.9 kDa protein. Q95LL4 46% 905 1174 HPIBO15 1310868 443 WUblastx.64 (Q9CQS3) 1110018M03RIK PROTEIN. Q9CQS3 93% 128 757 HPIBO15 590741 807 WUblastx.64 (Q9CQS3) 1110018M03RIK PROTEIN. Q9CQS3 88% 127 402 95% 507 722 97% 401 508 HPJCL22 1146674 445 WUblastx.64 (Q9GKV3) HYPOTHETICAL 41.8 KDA PROTEIN. Q9GKV3 92% 1540 2508 75% 2701 2823 HPJCL22 1034817 811 blastx.2 cDNA EST EMBL: M89462 comes from this gene; cDNA EST 1 1 emb|CAA94301.1| 44% 534 896 yk349d7.5 comes from this gene; cDNA EST yk358b9.5 comes from 29% 94 345 this HPJCL22 1046434 812 blastx.2 (AK000385) unnamed protein product [Homo sapiens] dbj|BAA91131.1| 71% 705 568 66% 743 702 HPJCW04 589969 446 WUblastx.64 (Q9N083) UNNAMED PORTEIN PRODUCT. Q9N083 45% 1275 1144 54% 1450 1265 HPMAI22 635491 448 WUblastx.64 (Q9CX19) 9430073N08RIK PROTEIN. Q9CX19 65% 147 629 HPQAC69 396804 451 WUblastx.64 (O75592) PROTEIN ASSOCIATED WITH MYC. O75592 100% 202 297 28% 76 189 100% 3 200 HPRBC80 829136 452 HMMER 2.1.1 PFAM: Protein phosphatase 2C PF00481 336.4 157 957 WUblastx.64 (Q9HAY8) SER/THR PROTEIN PHOSPHATASE TYPE 2C BETA 2 Q9HAY8 97% 94 1254 ISOFORM (PROTEIN HPRSB76 526310 453 WUblastx.64 (AAK33100) Aminophospholipid-transporting ATPase. AAK33100 38% 2 364 92% 112 570 HPWDJ42 722246 457 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 64% 1100 1026 67% 1332 1102 HPZAB47 585702 458 WUblastx.64 hypothetical protein 3 - human pir|E41925|E41925 34% 1132 884 55% 1296 1183 HRAAB15 658717 459 WUblastx.64 (Q9BVS2) UNKNOWN (PROTEIN FOR IMAGE: 3451448) Q9BVS2 40% 14 511 (FRAGMENT). HRABA80 882176 460 WUblastx.64 (Q9HA75) CDNA FLJ12122 FIS, CLONE MAMMA1000129. Q9HA75 63% 221 310 68% 325 459 HRABA80 588460 822 WUblastx.64 (Q9HA75) CDNA FLJ12122 FIS, CLONE MAMMA1000129. Q9HA75 63% 633 665 48% 130 357 92% 233 493 HRACD15 871221 461 WUblastx.64 (AAH08084) Hypothetical 50.4 kDa protein. AAH08084 98% 1452 253 HRACD80 1309774 462 WUblastx.64 (CAC37630) Fibulin-6 (Fragment). CAC37630 44% 700 1866 36% 37 1446 45% 1282 1920 42% 1291 1584 47% 1291 1530 HRACD80 882163 824 HMMER 2.1.1 PFAM: EGF-like domain PF00008 64.3 1337 1441 WUblastx.64 (CAC37630) Fibulin-6 (Fragment). CAC37630 44% 695 1861 37% 32 1441 45% 1277 1915 42% 1286 1579 47% 1286 1525 28% 1839 1913 33% 285 440 HRACD80 740762 825 blastx.2 (AF135253) fibulin-2 [Mus musculus] gb|AAD34456.1| 33% 898 1581 31% 1279 1893 45% 1279 1608 49% 681 911 44% 898 1125 30% 901 1605 43% 928 1149 31% 802 1137 43% 699 899 35% 690 998 41% 928 1119 42% 699 911 41% 699 908 48% 699 890 43% 699 914 33% 1339 1533 36% 898 1137 30% 540 995 33% 699 893 37% 699 890 43% 777 899 46% 925 1002 28% 898 1065 33% 493 609 26% 576 890 31% 952 1125 27% 807 1061 36% 943 1041 24% 699 869 23% 1057 1218 70% 1066 1095 35% 928 1053 HRDDV47 637650 463 WUblastx.64 (Q9VXD6) CG9723 PROTEIN. Q9VXD6 27% 83 964 HRDFD27 567004 464 WUblastx.64 (Q9N032) UNNAMED PROTEIN PRODUCT. Q9N032 47% 679 476 HSATR82 531973 466 WUblastx.64 (Q9UI58) PRO0483 PROTEIN. Q9UI58 80% 678 707 76% 605 682 HSAUL82 490879 468 WUblastx.64 (Q9BE22) HYPOTHETICAL 13.4 KDA PROTEIN. Q9BE22 63% 546 701 HSAVH65 545459 470 WUblastx.64 (Q9CZR4) 2700018N07RIK PROTEIN. Q9CZR4 92% 23 403 HSAVK10 561435 471 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 70% 1055 933 63% 1218 1030 HSAWZ41 580872 472 WUblastx.64 (Q9H387) PRO2550. Q9H387 81% 1386 1102 HSAXA83 545051 473 WUblastx.64 (Q9NRX6) PROTEIN X 013. Q9NRX6 100% 92 313 HSDAJ46 692358 475 HMMER 2.1.1 PFAM: Eukaryotic-type carbonic anhydrase PF00194 163.5 362 793 WUblastx.64 (Q9ULX7) CARBONIC ANHYDRASE XIV PRECURSOR (EC CAHE_HUMAN 99% 299 796 4.2.1.1) (CAR 98% 791 1084 HSDEK49 625998 827 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 18.7 225 470 WUblastx.64 (Q9Y279) Z39IG PROTEIN PRECURSOR. Q9Y279 88% 444 1040 99% 126 542 HSDEZ20 1352287 478 blastx.14 probable voltage-activated cation channel - rat pir|T17101|T17101 89% 4 336 60% 705 734 HSDEZ20 704101 828 WUblastx.64 probable voltage-activated cation channel - rat pir|T17101|T17101 89% 9 335 HSDJA15 795252 479 WUblastx.64 (Q9BZW5) TRANSMEMBRANE 6 SUPERFAMILY MEMBER 1. Q9BZW5 99% 4 702 HSDSE75 545057 481 WUblastx.64 (O60245) PCDH7 (BH-PCDH)A. O60245 100% 10 702 HSFAM31 552789 482 WUblastx.64 (Q9H728) CDNA: FLJ21463 FIS, CLONE COL04765. Q9H728 63% 868 836 63% 848 717 68% 741 589 HSHAX21 612823 483 WUblastx.64 (Q9NV22) CDNA FLJ10983 FIS, CLONE PLACE1001781, WEAKLY Q9NV22 99% 5 598 SIMILAR TO PRO HSIAS17 514183 830 WUblastx.64 (Q9H6H4) CDNA: FLJ22277 FIS, CLONE HRC03740. Q9H6H4 100% 108 362 88% 350 877 HSIDX71 1033671 485 WUblastx.64 (AAK55521) PRO0764. AAK55521 59% 1829 1764 65% 1786 1526 HSKDA27 1074734 832 WUblastx.64 (Q9CRM1) 2610001E17RIK PROTEIN (FRAGMENT). Q9CRM1 70% 793 1701 60% 1686 1784 23% 1604 1741 HSKDA27 872570 833 blastx.2 (AK020169) putative [Mus musculus] dbj|BAB32018.1| 47% 666 1562 HSSDX51 566879 495 WUblastx.64 (Q9NQ80) ASPIC PRECURSOR. Q9NQ80 83% 15 368 40% 301 399 72% 10 69 41% 174 266 32% 78 251 26% 99 257 92% 323 1105 HSSGD52 845666 839 WUblastx.64 (Q96FI8) Unknown (protein for MGC: 9160). Q96FI8 100% 338 2155 HSSJC35 1306937 498 WUblastx.64 (Q9H400) DJ583P15.4.1 (NOVEL PROTEIN (TRANSLATION OF Q9H400 81% 62 946 CDNA FLJ20406 (E HSSJC35 745409 840 WUblastx.64 (Q9H400) DJ583P15.4.1 (NOVEL PROTEIN (TRANSLATION OF Q9H400 100% 55 939 CDNA FLJ20406 (E HSUBW09 413246 500 WUblastx.64 (Q95LL0) Hypothetical 11.3 kDa protein. Q95LL0 73% 589 633 77% 327 611 HSVBU91 596868 502 WUblastx.64 cytoplasmic linker protein CLIP-115 - rat pir|T42734|T42734 85% 356 171 HSXCG83 944388 503 WUblastx.64 (Q9H7F4) CDNA: FLJ20979 FIS, CLONE ADSU01938. Q9H7F4 99% 101 901 HSXCG83 830673 842 blastx.2 (AL117204) Y116A8C.9 [Caenorhabditis elegans] emb|CAB55145.1| 36% 10 657 HSYAV50 847358 506 HMMER 2.1.1 PFAM: Leucine Rich Repeat PF00560 97.9 383 454 WUblastx.64 (Q96CX1) Similar to RIKEN cDNA 2610528G05 gene (Fragment). Q96CX1 96% 371 2170 HSYAZ63 1177537 509 WUblastx.64 (Q9Y613) FH1/FH2 DOMAINS-CONTAINING PROTEIN (FORMIN FHOS_HUMAN 98% 478 1713 HOMOLOG 96% 2573 2941 93% 2101 2514 55% 272 544 33% 790 933 33% 2030 2119 92% 3007 3090 28% 1015 1458 28% 289 654 42% 608 670 56% 2005 2052 41% 2220 2321 41% 2142 2255 37% 2098 2184 36% 2916 3005 42% 2913 2990 33% 2946 3026 69% 1756 1794 HSYAZ63 862063 848 WUblastx.64 (Q9Y613) FH1/FH2 DOMAINS-CONTAINING PROTEIN (FORMIN FHOS_HUMAN 78% 458 871 HOMOLOG 33% 387 476 92% 1364 1447 100% 14 70 96% 930 1298 69% 113 151 47% 558 620 36% 561 707 52% 362 418 32% 455 601 HSYBG37 1056317 510 WUblastx.64 hypothetical protein c316G12.3 [imported] - human pir|T45062|T45062 100% 122 961 HSYBG37 581098 849 WUblastx.64 hypothetical protein c316G12.3 [imported] - human pir|T45062|T45062 100% 48 962 HSZAF47 456551 850 HMMER 2.1.1 PFAM: Collagen triple helix repeat (20 copies) PF01391 54.4 299 478 WUblastx.64 (Q9BXJ2) COMPLEMENT-C1Q TUMOR NECROSIS FACTOR- Q9BXJ2 88% 500 976 RELATED PROTEIN. 62% 107 397 58% 344 394 50% 344 397 57% 353 394 HT3SF53 884170 512 WUblastx.64 (Q9H5B4) DJ470L14.2.1 (STAUFEN (RNA BINDING PROTEIN) Q9H5B4 100% 312 533 ISOFORM 1). HT5GJ57 1299921 513 WUblastx.64 (Q9GZY6) CDNA FLJ11237 FIS, CLONE PLACE1008531 Q9GZY6 89% 105 833 (WBSCR5) (WBSCR15 PROT HT5GJ57 740767 851 WUblastx.64 (Q9NZY9) HSPC046. Q9NZY9 90% 754 1002 70% 122 799 HTADX17 753289 514 WUblastx.64 (Q96A28) CD84-H1 (CD2 FAMILY 10). Q96A28 93% 92 412 79% 408 959 HTADX17 457172 852 WUblastx.64 (Q96A28) CD84-H1 (CD2 FAMILY 10). Q96A28 78% 490 585 97% 548 952 99% 84 488 HTDAF28 396835 515 WUblastx.64 (Q9BX79) STRA6 ISOFORM 1. Q9BX79 98% 17 298 HTEAF65 866485 516 WUblastx.64 (Q9DAC0) 1700013O04RIK PROTEIN. Q9DAC0 44% 9 287 HTEBI28 462221 517 WUblastx.64 (Q95LI0) Epididymis-specific protein ESP13.6. Q95LI0 46% 43 231 HTEDF80 587326 518 WUblastx.64 (Q9NP89) HYPOTHETICAL 42.7 KDA PROTEIN (FRAGMENT). Q9NP89 100% 253 327 30% 1016 1135 100% 852 1073 75% 112 210 98% 698 856 91% 353 451 66% 450 863 HTEDY42 519372 853 HMMER 2.1.1 PFAM: SCP-like extracellular protein PF00188 20 −98 −193 WUblastx.64 (Q96L06) Similar to RIKEN cDNA 1700011E04 gene. Q96L06 100% 19 231 33% 576 719 94% 224 700 HTEGI42 908143 521 WUblastx.64 (AAH20905) Hypothetical 28.5 kDa protein. AAH20905 88% 41 796 HTEHR24 835894 522 WUblastx.64 (Q9HBV2) SPERM MEMBRANE ANTIGEN SMARC32. Q9HBV2 76% 84 959 HTEHR24 513039 858 WUblastx.64 (Q9HBV2) SPERM MEMBRANE ANTIGEN SMARC32. Q9HBV2 76% 41 529 100% 692 922 96% 514 693 HTEHU31 600394 523 WUblastx.64 (Q9NPE6) DJ309K20.2 (ACROSOMAL PROTEIN ACR55 (SIMILAR Q9NPE6 97% 16 1056 TO RAT SPERMAN HTEHU93 722254 524 WUblastx.64 (O60676) CYSTATIN-RELATED EPIDIDYMAL CRES_HUMAN 91% 188 613 SPERMATOGENIC PROTEIN HTEHU93 423009 859 HMMER 2.1.1 PFAM: Cystatin domain PF00031 31.7 35 −105 WUblastx.64 (O60676) CYSTATIN-RELATED EPIDIDYMAL CRES_HUMAN 100% 504 614 SPERMATOGENIC PROTEIN 78% 187 552 HTEJN13 658744 860 WUblastx.64 (Q9DAR9) 1700001D09RIK PROTEIN. Q9DAR9 60% 525 743 77% 163 516 HTEPG70 834931 529 WUblastx.64 (O75295) R27328_2. O75295 93% 23 268 HTGAU75 597467 530 WUblastx.64 (Q9NZX5) HSPC062. Q9NZX5 55% 502 672 72% 149 661 HTGEP89 410582 531 WUblastx.64 (Q9DAL9) 1700007K09RIK PROTEIN. Q9DAL9 44% 258 566 HTHBG43 919911 532 WUblastx.64 (Q9H387) PRO2550. Q9H387 83% 772 701 70% 702 571 HTHDJ94 693652 534 HMMER 2.1.1 PFAM: Oxidoreductase FAD/NAD-binding domain PF00175 160.3 552 896 WUblastx.64 (Q9UHQ9) NADH-CYTOCHROME B5 REDUCTASE ISOFORM. Q9UHQ9 95% 66 941 HTHDS25 772559 535 WUblastx.64 (Q9P1H3) PRO1438. Q9P1H3 66% 1045 911 HTJMA95 706618 536 HMMER 2.1.1 PFAM: Ammonium Transporter Family PF00909 62.1 533 691 WUblastx.64 (Q9UBD6) RH TYPE C GLYCOPROTEIN (TUMOR-RELATED Q9UBD6 98% 3 455 PROTEIN DRC2). 100% 449 1069 HTJML75 1040047 537 WUblastx.64 (Q9UJX6) ANAPHASE-PROMOTING COMPLEX SUBUNIT 2. Q9UJX6 94% 30 2495 HTJML75 873355 864 WUblastx.64 (Q9UJX6) ANAPHASE-PROMOTING COMPLEX SUBUNIT 2. Q9UJX6 78% 40 423 94% 423 1016 89% 911 2503 HTLBE23 902187 538 WUblastx.64 (Q96M29) CDNA FLJ32871 fis, clone TESTI2003914, weakly similar Q96M29 98% 176 838 to Tek 93% 840 980 81% 1112 1177 HTLFE42 460583 539 WUblastx.64 (Q9NSI0) PRED58 PROTEIN (FRAGMENT). Q9NSI0 99% 17 346 HTLFE57 791409 866 WUblastx.64 (Q9D7G6) 2310009N05RIK PROTEIN. Q9D7G6 90% 12 698 HTLFE57 608317 867 WUblastx.64 (Q9D7G6) 2310009N05RIK PROTEIN. Q9D7G6 84% 2 619 HTLGE31 1035130 541 WUblastx.64 (Q9NY64) GLUCOSE TRANSPORTER. Q9NY64 100% 3 92 HTLHY14 838460 542 WUblastx.64 (Q96L02) Hypothetical 24.5 kDa protein. Q96L02 99% 36 434 100% 528 773 HTLIT32 833906 543 WUblastx.64 (Q96QH1) NB1 Glycoprotein precursor. Q96QH1 32% 312 932 29% 330 1007 HTLIV19 1046341 544 WUblastx.64 (Q96LS9) CDNA FLJ25101 fis, clone CBR01328. Q96LS9 73% 193 315 HTODK73 526021 547 WUblastx.64 (Q9H8P2) CDNA FLJ13348 FIS, CLONE OVARC1002127, Q9H8P2 93% 404 448 WEAKLY SIMILAR TO SOD 100% 567 707 71% 433 474 43% 4 189 61% 418 519 80% 21 401 HTOHM15 1028538 551 WUblastx.64 (Q9NVL9) CDNA FLJ10649 FIS, CLONE NT2RP2005835, WEAKLY Q9NVL9 96% 1641 1718 SIMILAR TO SHP 100% 1507 1650 HTOHM15 848200 870 HMMER 2.1.1 PFAM: UBX domain PF00789 97.6 794 1033 HTOJA73 797108 554 WUblastx.64 (Q9H387) PRO2550. Q9H387 63% 1044 955 66% 1294 1046 HTOJK60 545067 555 WUblastx.64 (Q9HA67) CDNA FLJ12155 FIS, CLONE MAMMA1000472. Q9HA67 73% 745 644 78% 870 757 HTPBW79 1317835 556 WUblastx.64 (Q9CXR7) 3110023E09RIK PROTEIN. Q9CXR7 77% 172 813 92% 787 999 HTPBW79 581435 873 WUblastx.64 (Q96S93) Hypothetical 41.7 kDa protein. Q96S93 95% 302 1387 HTTDB46 812763 559 WUblastx.64 (Q9Y2C7) BUTYROPHILIN LIKE RECEPTOR. Q9Y2C7 70% 106 543 83% 727 762 59% 1007 1072 100% 1644 2180 HTTDB46 909573 875 HMMER 2.1.1 PFAM: SPRY domain PF00622 65.9 −956 −1276 HTWCT03 429618 560 WUblastx.64 (O95014) WUGSC:H_DJ0855D21.2 PROTEIN. O95014 82% 1488 1592 HTWDF76 714344 561 WUblastx.64 (Q9BTF2) REC8P, A MEIOTIC RECOMBINATION AND SISTER Q9BTF2 100% 792 875 CHROMATID COHESION 92% 370 510 27% 7 498 35% 179 238 37% 379 525 79% 542 688 70% 179 280 76% 4 192 HTXDW56 695765 565 WUblastx.64 (Q96A54) Similar to CGI-45 protein (Hypothetical 42.6 kDa protein). Q96A54 99% 7 819 HTXFL30 620001 566 WUblastx.64 (Q96KR5) Leishmanolysin-like peptidase, variant 2 (EC 3.4.24.36). Q96KR5 98% 305 1990 100% 30 68 100% 213 299 100% 68 94 HTXKP61 824083 567 WUblastx.64 (Q9H0S8) HYPOTHETICAL 53.0 KDA PROTEIN. Q9H0S8 98% 3 1124 HUDBZ89 562791 877 WUblastx.64 (Q9VH80) CG16908 PROTEIN. Q9VH80 22% 7 327 33% 330 641 HUFEF62 645101 570 WUblastx.64 hypothetical L1 protein (third intron of gene TS) - human pir|JU0033|JU0033 81% 355 308 84% 314 12 HUKAH51 1300737 880 WUblastx.64 (Q9ES75) PROLINE-RICH ACIDIC PROTEIN. Q9ES75 50% 144 563 HUKAH51 603538 881 WUblastx.64 (Q96NZ9) Proline-rich acidic protein. Q96NZ9 100% 462 479 93% 55 462 HUKBT29 694590 572 WUblastx.64 (Q96AA2) Obscurin. Q96AA2 82% 131 1300 30% 520 597 33% 500 571 29% 152 370 100% 1039 1338 34% 597 710 28% 134 316 HUSAT94 606599 573 WUblastx.64 (Q96LS9) CDNA FLJ25101 fis, clone CBR01328. Q96LS9 60% 1865 1752 73% 1752 1573 HUSBA88 895435 574 HMMER 2.1.1 PFAM: Glycosyl hydrolase family 47 PF01532 694 783 2102 WUblastx.64 (Q9UKM7) ALPHA 1,2-MANNOSIDASE. Q9UKM7 100% 18 2114 HUSIG64 566762 575 WUblastx.64 (O60763) GENERAL VESICULAR TRANSPORT FACTOR P115 VDP_HUMAN 99% 9 1010 (TRANSCYTO HUSXS50 883176 882 WUblastx.64 (AAH08361) F-box only protein 7. AAH08361 99% 281 1069 42% 1566 1622 100% 1067 1666 HUSXS50 655372 883 WUblastx.64 (AAH08361) F-box only protein 7. AAH08361 77% 1 459 26% 43 219 100% 317 700 HWAAD63 838626 577 HMMER 2.1.1 PFAM: Sodium/calcium exchanger protein PF01699 62.8 346 453 WUblastx.64 (Q9HC58) SODIUM/CALCIUM EXCHANGER NCKX3. Q9HC58 65% 229 813 HWAAD63 833089 884 HMMER 2.1.1 PFAM: Sodium/calcium exchanger protein PF01699 37.8 346 453 blastx.2 (AF177984) potassium-dependent sodium-calcium exchanger NCKX1 gb|AAF25808.1| 45% 217 453 [Gallus gallus] AF177984_1 41% 533 793 45% 453 596 31% 319 453 HWAAD63 793875 885 HMMER 2.1.1 PFAM: Sodium/calcium exchanger protein PF01699 113.7 336 773 blastx.2 (AF025664) Na—Ca + K exchanger [Bos taurus] gb|AAB88884.1| 43% 207 785 HWABY10 768334 579 WUblastx.64 (Q96AW1) Hypothetical 19.2 kDa protein. Q96AW1 100% 165 665 HWBAO62 838164 581 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 27.9 202 402 WUblastx.64 (Q14288) HYPOTHETICAL PROTEIN (FRAGMENT). Q14288 45% 1331 1618 66% 1158 1334 62% 1847 1894 55% 1594 1839 HWBAR14 1107118 582 WUblastx.64 (AAK54386) Prostein. AAK54386 80% 1737 2426 84% 290 1033 HWBAR88 836469 583 WUblastx.64 (Q9Y2C2) DERMATAN/CHONDROITIN SULFATE 2- Q9Y2C2 83% 958 1050 SULFOTRANSFERASE. 64% 107 241 88% 215 982 HWBCB89 1093347 584 WUblastx.64 (BAB55294) CDNA FLJ14777 fis, clone NT2RP4000259, w BAB55294 100% 94 576 HWBCB89 886210 890 HMMER 2.1.1 PFAM: Glutathione peroxidases PF00255 170.2 104 433 WUblastx.64 (BAB55294) CDNA FLJ14777 fis, clone NT2RP4000259, w BAB55294 100% 35 595 HWBCP79 846382 585 WUblastx.64 (AAH20829) Hypothetical 6.2 kDa protein. AAH20829 78% 134 93 78% 72 16 HWBCP79 646977 891 WUblastx.64 (Q96MM0) CDNA FLJ32172 fis, clone PLACE6000555. Q96MM0 27% 330 133 85% 148 68 HWBEM18 949402 587 WUblastx.64 nuclear pore protein gp210 precursor - rat pir|S04921|S04921 83% 159 5735 HWBEM18 906580 893 blastx.2 gp210 (AA 1-1886) [Rattus norvegicus] emb|CAA68759.1| 87% 131 2629 79% 2626 3570 31% 2595 2732 HWBEM18 877573 894 WUblastx.64 hypothetical protein DKFZp434P1650.1 - human (fragment) pir|T17289|T17289 96% 205 1494 HWBFE57 907063 588 WUblastx.64 (Q9NR73) MACROPHAGE ABC TRANSPORTER. Q9NR73 78% 206 1048 HWDAH38 1028519 590 WUblastx.64 (Q9NX85) CDNA FLJ20378 FIS, CLONE KAIA0536. Q9NX85 69% 1113 1250 63% 979 1119 81% 947 979 52% 1534 1340 60% 1602 1528 HWHGP71 995431 591 HMMER 2.1.1 PFAM: 7 transmembrane receptor (rhodopsin family) PF00001 31.2 389 766 WUblastx.64 leukotriene B4 receptor 2, BLTR2 - human pir|JC7356|JC7356 94% 101 766 35% 487 591 47% 434 484 84% 715 1020 HWHGP71 839250 899 blastx.2 (AJ278605) leukotriene B4 receptor 2 [Homo sapiens] emb|CAB96134.1| 77% 106 465 100% 555 770 58% 776 1036 HWHGQ49 636080 900 WUblastx.64 (Q9YSB4) ANDROGEN INDUCED PROTEIN. Q9Y5B4 99% 42 755 HWHGU54 695695 593 HMMER 2.1.1 PFAM: Serpins (serine protease inhibitors) PF00079 501.1 277 1377 WUblastx.64 (Q9CQ32) 4632419J12RIK PROTEIN. Q9CQ32 61% 145 1383 HWHGZ51 886212 594 WUblastx.64 (Q9UJ74) HYPOTHETICAL 36.0 KDA PROTEIN (C4.4A PROTEIN). Q9UJ74 86% 33 1022 HWHHL34 805642 595 WUblastx.64 (O75915) JWA PROTEIN (HSPC127) (VITAMIN A RESPONSIVE, O75915 100% 131 694 CYTOSKELETON RE HWHHL34 801943 901 blastx.2 (AF070523) JWA protein [Homo sapiens] gb|AAC64360.1| 92% 53 613 HWHQS55 762842 596 HMMER 2.1.1 PFAM: Cadherin domain PF00028 224.2 907 1182 WUblastx.64 (AAK51616) Protocadherin-beta10. AAK51616 99% 169 1800 30% 535 1785 80% 1817 2563 28% 520 1632 HWLEV32 1032602 597 WUblastx.64 (Q63778) HYPOTHETICAL 43.7 KDA PROTEIN. Q63778 54% 142 20 40% 400 239 38% 419 342 48% 684 535 HWLEV32 846351 905 WUblastx.64 (Q9W6Q6) OSTEOBLAST 6D12C PROTEIN. Q9W6Q6 88% 143 421 HWLIH65 793713 598 HMMER 2.1.1 PFAM: Integral membrane protein PF01940 49.3 147 455 WUblastx.64 (AAH08596) Unknown (protein for MGC: 16985). AAH08596 98% 81 623 HYAAJ71 826754 599 WUblastx.64 (Q9NX17) CDNA FLJ20489 FIS, CLONE KAT08285. Q9NX17 62% 1147 1464 HAPSA79 846517 602 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 67 924 1130 WUblastx.64 (Q9BX67) JUNCTIONAL ADHESION MOLECULE 3 PRECURSOR. Q9BX67 100% 468 1397 HAPSA79 887467 906 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 67 924 1130 blastx.2 (AF356518) junctional adhesion molecule 3 precursor [Homo sapiens] gb|AAK27221.1| 92% 540 1397 AF356518_1 HAPSA79 878627 907 HMMER 2.1.1 PFAM: Immunoglobulin domain PF00047 67 924 1130 WUblastx.64 (Q9BX67) JUNCTIONAL ADHESION MOLECULE 3 PRECURSOR. Q9BX67 100% 468 1397

RACE Protocol For Recovery of Full-Length Genes

Partial cDNA clones can be made full-length by utilizing the rapid amplification of cDNA ends (RACE) procedure described in Frohman, M. A., et al., Proc. Nat'l. Acad. Sci. USA, 85:8998-9002 (1988). A cDNA clone missing either the 5′ or 3′ end can be reconstructed to include the absent base pairs extending to the translational start or stop codon, respectively. In some cases, cDNAs are missing the start codon of translation, therefor. The following briefly describes a modification of this original 5′ RACE procedure. Poly A+ or total RNA is reverse transcribed with Superscript II (Gibco/BRL) and an antisense or complementary primer specific to the cDNA sequence. The primer is removed from the reaction with a Microcon Concentrator (Amicon). The first-strand cDNA is then tailed with dATP and terminal deoxynucleotide transferase (Gibco/BRL). Thus, an anchor sequence is produced which is needed for PCR amplification. The second strand is synthesized from the dA-tail in PCR buffer, Taq DNA polymerase (Perkin-Elmer Cetus), an oligo-dT primer containing three adjacent restriction sites (XhoI, SalI and ClaI) at the 5′ end and a primer containing just these restriction sites. This double-stranded cDNA is PCR amplified for 40 cycles with the same primers as well as a nested cDNA-specific antisense primer. The PCR products are size-separated on an ethidium bromide-agarose gel and the region of gel containing cDNA products the predicted size of missing protein-coding DNA is removed. cDNA is purified from the agarose with the Magic PCR Prep kit (PROMEGA™), restriction digested with XhoI or SalI, and ligated to a plasmid such as pBLUESCRIPT™ SKII (STRATAGENE™) at XhoI and EcoRV sites. This DNA is transformed into bacteria and the plasmid clones sequenced to identify the correct protein-coding inserts. Correct 5′ ends are confirmed by comparing this sequence with the putatively identified homologue and overlap with the partial cDNA clone. Similar methods known in the art and/or commercial kits are used to amplify and recover 3′ ends.

Several quality-controlled kits are commercially available for purchase. Similar reagents and methods to those above are supplied in kit form from Gibco/BRL for both 5′ and 3′ RACE for recovery of full length genes. A second kit is available from CLONTECH™ which is a modification of a related technique, SLIC (single-stranded ligation to single-stranded cDNA), developed by Dumas et al., Nucleic Acids Res., 19:5227-32 (1991). The major differences in procedure are that the RNA is alkaline hydrolyzed after reverse transcription and RNA ligase is used to join a restriction site-containing anchor primer to the first-strand cDNA. This obviates the necessity for the dA-tailing reaction which results in a polyT stretch that is difficult to sequence past.

An alternative to generating 5′ or 3′ cDNA from RNA is to use cDNA library double-stranded DNA. An asymmetric PCR-amplified antisense cDNA strand is synthesized with an antisense cDNA-specific primer and a plasmid-anchored primer. These primers are removed and a symmetric PCR reaction is performed with a nested cDNA-specific antisense primer and the plasmid-anchored primer.

RNA Ligase Protocol for Generating the 5′ or 3′ End Sequences to Obtain Full Length Genes

Once a gene of interest is identified, several methods are available for the identification of the 5′ or 3′ portions of the gene which may not be present in the original cDNA plasmid. These methods include, but are not limited to, filter probing, clone enrichment using specific probes and protocols similar and identical to 5′ and 3′ RACE. While the full length gene may be present in the library and can be identified by probing, a useful method for generating the 5′ or 3′ end is to use the existing sequence information from the original cDNA to generate the missing information. A method similar to 5′ RACE is available for generating the missing 5′ end of a desired full-length gene. (This method was published by Fromont-Racine et al., Nucleic Acids Res., 21(7):1683-1684 (1993)). Briefly, a specific RNA oligonucleotide is ligated to the 5′ ends of a population of RNA presumably containing full-length gene RNA transcript and a primer set containing a primer specific to the ligated RNA oligonucleotide and a primer specific to a known sequence of the gene of interest, is used to PCR amplify the 5′ portion of the desired full length gene which may then be sequenced and used to generate the full length gene. This method starts with total RNA isolated from the desired source, poly A RNA may be used but is not a prerequisite for this procedure. The RNA preparation may then be treated with phosphatase if necessary to eliminate 5′ phosphate groups on degraded or damaged RNA which may interfere with the later RNA ligase step. The phosphatase if used is then inactivated and the RNA is treated with tobacco acid pyrophosphatase in order to remove the cap structure present at the 5′ ends of messenger RNAs. This reaction leaves a 5′ phosphate group at the 5′ end of the cap cleaved RNA which can then be ligated to an RNA oligonucleotide using T4 RNA ligase. This modified RNA preparation can then be used as a template for first strand cDNA synthesis using a gene specific oligonucleotide. The first strand synthesis—reaction can then be used as a template for PCR amplification of the desired 5′ end using a primer specific to the ligated RNA oligonucleotide and a primer specific to the known sequence of the gene of interest. The resultant product is then sequenced and analyzed to confirm that the 5′ end sequence belongs to the relevant gene.

The present invention also relates to vectors or plasmids which include such DNA sequences, as well as the use of the DNA sequences. The material deposited with the ATCC™ (e.g., as described in columns 2 and 3 of Table 1A, and/or as set forth in Table 1B, Table 6, or Table 7) is a mixture of cDNA clones derived from a variety of human tissue and cloned in either a plasmid vector or a phage vector, as described, for example, in Table 1A and Table 7. These deposits are referred to as “the deposits” herein. The tissues from which some of the clones were derived are listed in Table 7, and the vector in which the corresponding cDNA is contained is also indicated in Table 7. The deposited material includes cDNA clones corresponding to SEQ ID NO:X described, for example, in Table 1A and/or Table 1B (ATCC™ Deposit No:Z). A clone which is isolatable from the ATCC™ Deposits by use of a sequence listed as SEQ ID NO:X, may include the entire coding region of a human gene or in other cases such clone may include a substantial portion of the coding region of a human gene. Furthermore, although the sequence listing may in some instances list only a portion of the DNA sequence in a clone included in the ATCC™ Deposits, it is well within the ability of one skilled in the art to sequence the DNA included in a clone contained in the ATCC™ Deposits by use of a sequence (or portion thereof) described in, for example Tables 1A and/or Table 1B or Table 2, by procedures hereinafter further described, and others apparent to those skilled in the art.

Also provided in Table 1A and Table 7 is the name of the vector which contains the cDNA clone. Each vector is routinely used in the art. The following additional information is provided for convenience.

Vectors Lambda Zap (U.S. Pat. Nos. 5,128,256 and 5,286,636), Uni-Zap XR (U.S. Pat. Nos. 5,128,256 and 5,286,636), Zap Express (U.S. Pat. Nos. 5,128,256 and 5,286,636), pBLUESCRIPT™ (pBS) (Short, J. M. et al., Nucleic Acids Res. 16:7583-7600 (1988); Alting-Mees, M. A. and Short, J. M., Nucleic Acids Res. 17:9494 (1989)) and pBK (Alting-Mees, M. A. et al., Strategies 5:58-61 (1992)) are commercially available from STRATAGENE™ Cloning Systems, Inc., 11011 N. Torrey Pines Road, La Jolla, Calif., 92037. pBS contains an ampicillin resistance gene and pBK contains a neomycin resistance gene. Phagemid pBS may be excised from the Lambda Zap and Uni-Zap XR vectors, and phagemid pBK may be excised from the Zap Express vector. Both phagemids may be transformed into E. coli strain XL-1 Blue, also available from STRATAGENE™.

Vectors pSport1, pCMVSport 1.0, pCMVSport 2.0 and pCMVSport 3.0, were obtained from Life Technologies, Inc., P.O. Box 6009, Gaithersburg, Md. 20897. All Sport vectors contain an ampicillin resistance gene and may be transformed into E. coli strain DH10B, also available from Life Technologies. See, for instance, Gruber, C. E., et al., Focus 15:59-(1993). Vector lafmid BA (Bento Soares, Columbia University, New York, N.Y.) contains an ampicillin resistance gene and can be transformed into E. coli strain XL-1 Blue. Vector pCR®2.1, which is available from Invitrogen, 1600 Faraday Avenue, Carlsbad, Calif. 92008, contains an ampicillin resistance gene and may be transformed into E. coli strain DH10B, available from Life Technologies. See, for instance, Clark, J. M., Nuc. Acids Res. 16:9677-9686 (1988) and Mead, D. et al., Bio/Technology 9: (1991).

The present invention also relates to the genes corresponding to SEQ ID NO:X, SEQ ID NO:Y, and/or the deposited clone (ATCC™ Deposit No:Z). The corresponding gene can be isolated in accordance with known methods using the sequence information disclosed herein. Such methods include preparing probes or primers from the disclosed sequence and identifying or amplifying the corresponding gene from appropriate sources of genomic material.

Also provided in the present invention are allelic variants, orthologs, and/or species homologs. Procedures known in the art can be used to obtain full-length genes, allelic variants, splice variants, full-length coding portions, orthologs, and/or species homologs of genes corresponding to SEQ ID NO:X or the complement thereof, polypeptides encoded by genes corresponding to SEQ ID NO:X or the complement thereof, and/or the cDNA contained in ATCC™ Deposit No:Z, using information from the sequences disclosed herein or the clones deposited with the ATCC™. For example, allelic variants and/or species homologs may be isolated and identified by making suitable probes or primers from the sequences provided herein and screening a suitable nucleic acid source for allelic variants and/or the desired homologue.

The polypeptides of the invention can be prepared in any suitable manner. Such polypeptides include isolated naturally occurring polypeptides, recombinantly produced polypeptides, synthetically produced polypeptides, or polypeptides produced by a combination of these methods. Means for preparing such polypeptides are well understood in the art.

The polypeptides may be in the form of the secreted protein, including the mature form, or may be a part of a larger protein, such as a fusion protein (see below). It is often advantageous to include an additional amino acid sequence which contains secretory or leader sequences, pro-sequences, sequences which aid in purification, such as multiple histidine residues, or an additional sequence for stability during recombinant production.

The polypeptides of the present invention are preferably provided in an isolated form, and preferably are substantially purified. A recombinantly produced version of a polypeptide, including the secreted polypeptide, can be substantially purified using techniques described herein or otherwise known in the art, such as, for example, by the one-step method described in Smith and Johnson, Gene 67:31-40 (1988). Polypeptides of the invention also can be purified from natural, synthetic or recombinant sources using techniques described herein or otherwise known in the art, such as, for example, antibodies of the invention raised against the polypeptides of the present invention in methods which are well known in the art.

The present invention provides a polynucleotide comprising, or alternatively consisting of, the nucleic acid sequence of SEQ ID NO:X, and/or the cDNA sequence contained in ATCC™ Deposit No:Z. The present invention also provides a polypeptide comprising, or alternatively, consisting of, the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X or a complement thereof, a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z, and/or the polypeptide sequence encoded by a nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1C. Polynucleotides encoding a polypeptide comprising, or alternatively consisting of the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X, a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z, and/or a polypeptide sequence encoded by a nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1C are also encompassed by the invention. The present invention further encompasses a polynucleotide comprising, or alternatively consisting of, the complement of the nucleic acid sequence of SEQ ID NO:X, a nucleic acid sequence encoding a polypeptide encoded by the complement of the nucleic acid sequence of SEQ ID NO:X, and/or the cDNA contained in ATCC™ Deposit No:Z.

Moreover, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in Table 1C column 6, or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in Table 1C column 6, or any combination thereof. In further embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in Table 1C, column 6, and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1C, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in Table 1C, column 6, and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in Table 1C, column 6, and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

Further, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1C which correspond to the same Clone ID (see Table 1C, column 1), or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in column 6 of Table 1C which correspond to the same Clone ID (see Table 1C, column 1), or any combination thereof. In further embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1C which correspond to the same Clone ID (see Table 1C, column 1) and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1C, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1C which correspond to the same Clone ID (see Table 1C, column 1) and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1C which correspond to the same Clone ID (see Table 1C, column 1) and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

Further, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1C which correspond to the same contig sequence identifier SEQ ID NO:X (see Table 1C, column 2), or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in column 6 of Table 1C which correspond to the same contig sequence identifier SEQ ID NO:X (see Table 1C, column 2), or any combination thereof. In further embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1C which correspond to the same contig sequence identifier SEQ ID NO:X (see Table 1C, column 2) and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1C, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1C which correspond to the same contig sequence identifier SEQ ID NO:X (see Table 1C, column 2) and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1C which correspond to the same contig sequence identifier SEQ ID NO:X (see Table 1C, column 2) and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (See Table 1C, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

Moreover, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in the same row of Table 1C column 6, or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in the same row of Table 1C column 6, or any combination thereof. In preferred embodiments, the polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in the same row of Table 1C column 6, wherein sequentially delineated sequences in the table (i.e. corresponding to those exons located closest to each other) are directly contiguous in a 5′ to 3′ orientation. In further embodiments, above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in the same row of Table 1C, column 6, and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1C, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in the same row of Table 1C, column 6, and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in the same row of Table 1C, column 6, and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1C, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1C, column 2) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1C which correspond to the same Clone ID (see Table 1C, column 1), and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A, Table 1B, or Table 1C) or fragments or variants thereof. In preferred embodiments, the delineated sequence(s) and polynucleotide sequence of SEQ ID NO:X correspond to the same Clone ID. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in the same row of column 6 of Table 1C, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A, Table 1B, or Table 1C) or fragments or variants thereof. In preferred embodiments, the delineated sequence(s) and polynucleotide sequence of SEQ ID NO:X correspond to the same row of column 6 of Table 1C. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of the sequence of SEQ ID NO:X are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X are directly contiguous Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of the sequence of SEQ ID NO:X and the 5′ 10 polynucleotides of the sequence of one of the sequences delineated in column 6 of Table 1C are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X and the 5′ 10 polynucleotides of the sequence of one of the sequences delineated in column 6 of Table 1C are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides, are also encompassed by the invention.

In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of another sequence in column 6 are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of another sequence in column 6 corresponding to the same Clone ID (see Table 1C, column 1) are directly contiguous. Nucleic acids which hybridize to the complement of these 20 lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one sequence in column 6 corresponding to the same contig sequence identifier SEQ ID NO:X (see Table 1C, column 2) are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of another sequence in column 6 corresponding to the same row are directly contiguous. In preferred embodiments, the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C is directly contiguous with the 5′ 10 polynucleotides of the next sequential exon delineated in Table 1C, column 6 Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

Table 3

Many polynucleotide sequences, such as EST sequences, are publicly available and accessible through sequence databases and may have been publicly available prior to conception of the present invention. Preferably, such related polynucleotides are specifically excluded from the scope of the present invention. Accordingly, for each contig sequence (SEQ ID NO:X) listed in the fifth column of Table 1A and/or the fourth column of Table 1B, preferably excluded are one or more polynucleotides comprising a nucleotide sequence described by the general formula of a-b, where a is any integer between 1 and the final nucleotide minus 15 of SEQ ID NO:X, b is an integer of 15 to the final nucleotide of SEQ ID NO:X, where both a and b correspond to the positions of nucleotide residues shown in SEQ ID NO:X, and where b is greater than or equal to a +14. More specifically, preferably excluded are one or more polynucleotides comprising a nucleotide sequence described by the general formula of a-b, where a and b are integers as defined in columns 4 and 5, respectively, of Table 3. In specific embodiments, the polynucleotides of the invention do not consist of at least one, two, three, four, five, ten, or more of the specific polynucleotide sequences referenced by the Genbank Accession No. as disclosed in column 6 of Table 3 (including for example, published sequence in connection with a particular BAC clone). In further embodiments, preferably excluded from the invention are the specific polynucleotide sequence(s) contained in the clones corresponding to at least one, two, three, four, five, ten, or more of the available material having the accession numbers identified in the sixth column of this Table (including for example, the actual sequence contained in an identified BAC clone). In no way is this listing meant to encompass all of the sequences which may be excluded by the general formula, it is just a representative example. All references available through these accessions are hereby incorporated by reference in their entirety. Table 3, from U.S. patent application Ser. No. 10/472,532, filed Sep. 20, 2003, is herein incorporated by reference.

Description of Table 4

Table 4 provides a key to the tissue/cell source identifier code disclosed in Table 1B.2, column 5. Column 1 provides the tissue/cell source identifier code disclosed in Table 1B.2, Column 5. Columns 2-5 provide a description of the tissue or cell source. Note that “Description” and “Tissue” sources (i.e. columns 2 and 3) having the prefix “a_” indicates organs, tissues, or cells derived from “adult” sources. Codes corresponding to diseased tissues are indicated in column 6 with the word “disease.” The use of the word “disease” in column 6 is non-limiting. The tissue or cell source may be specific (e.g. a neoplasm), or may be disease-associated (e.g., a tissue sample from a normal portion of a diseased organ). Furthermore, tissues and/or cells lacking the “disease” designation may still be derived from sources directly or indirectly involved in a disease state or disorder, and therefore may have a further utility in that disease state or disorder. In numerous cases where the tissue/cell source is a library, column 7 identifies the vector used to generate the library.

TABLE 4 Code Description Tissue AR022 a_Heart a_Heart AR023 a_Liver a_Liver AR024 a_mammary gland a_mammary gland AR025 a_Prostate a_Prostate AR026 a_small intestine a_small intestine AR027 a_Stomach a_Stomach AR028 Blood B cells Blood B cells AR029 Blood B cells activated Blood B cells activated AR030 Blood B cells resting Blood B cells resting AR031 Blood T cells activated Blood T cells activated AR032 Blood T cells resting Blood T cells resting AR033 brain brain AR034 breast breast AR035 breast cancer breast cancer AR036 Cell Line CAOV3 Cell Line CAOV3 AR037 cell line PA-1 cell line PA-1 AR038 cell line transformed cell line transformed AR039 colon colon AR040 colon (9808co65R) colon (9808co65R) AR041 colon (9809co15) colon (9809co15) AR042 colon cancer colon cancer AR043 colon cancer (9808co64R) colon cancer (9808co64R) AR044 colon cancer 9809co14 colon cancer 9809co14 AR050 Donor II B Cells 24 hrs Donor II B Cells 24 hrs AR051 Donor II B Cells 72 hrs Donor II B Cells 72 hrs AR052 Donor II B-Cells 24 hrs. Donor II B-Cells 24 hrs. AR053 Donor II B-Cells 72 hrs Donor II B-Cells 72 hrs AR054 Donor II Resting B Cells Donor II Resting B Cells AR055 Heart Heart AR056 Human Lung Human Lung (CLONTECH ™) (CLONTECH ™) AR057 Human Mammary Human Mammary (CLONTECH ™) (CLONTECH ™) AR058 Human Thymus Human Thymus (CLONTECH ™) (CLONTECH ™) AR059 Jurkat (unstimulated) Jurkat (unstimulated) AR060 Kidney Kidney AR061 Liver Liver AR062 Liver (CLONTECH ™) Liver (CLONTECH ™) AR063 Lymphocytes chronic Lymphocytes chronic lymphocytic leukaemia lymphocytic leukaemia AR064 Lymphocytes diffuse large B Lymphocytes diffuse large B cell lymphoma cell lymphoma AR065 Lymphocytes follicular Lymphocytes follicular lymphoma lymphoma AR066 normal breast normal breast AR067 Normal Ovarian (4004901) Normal Ovarian (4004901) AR068 Normal Ovary 9508G045 Normal Ovary 9508G045 AR069 Normal Ovary 9701G208 Normal Ovary 9701G208 AR070 Normal Ovary 9806G005 Normal Ovary 9806G005 AR071 Ovarian Cancer Ovarian Cancer AR072 Ovarian Cancer (9702G001) Ovarian Cancer (9702G001) AR073 Ovarian Cancer (9707G029) Ovarian Cancer (9707G029) AR074 Ovarian Cancer (9804G011) Ovarian Cancer (9804G011) AR075 Ovarian Cancer (9806G019) Ovarian Cancer (9806G019) AR076 Ovarian Cancer (9807G017) Ovarian Cancer (9807G017) AR077 Ovarian Cancer (9809G001) Ovarian Cancer (9809G001) AR078 ovarian cancer 15799 ovarian cancer 15799 AR079 Ovarian Cancer 17717AID Ovarian Cancer 17717AID AR080 Ovarian Cancer 4004664B1 Ovarian Cancer 4004664B1 AR081 Ovarian Cancer 4005315A1 Ovarian Cancer 4005315A1 AR082 ovarian cancer 94127303 ovarian cancer 94127303 AR083 Ovarian Cancer 96069304 Ovarian Cancer 96069304 AR084 Ovarian Cancer 9707G029 Ovarian Cancer 9707G029 AR085 Ovarian Cancer 9807G045 Ovarian Cancer 9807G045 AR086 ovarian cancer 9809G001 ovarian cancer 9809G001 AR087 Ovarian Cancer Ovarian Cancer 9905C032RC 9905C032RC AR088 Ovarian cancer 9907 C00 3rd Ovarian cancer 9907 C00 3rd AR089 Prostate Prostate AR090 Prostate (CLONTECH ™) Prostate (CLONTECH ™) AR091 prostate cancer prostate cancer AR092 prostate cancer #15176 prostate cancer #15176 AR093 prostate cancer #15509 prostate cancer #15509 AR094 prostate cancer #15673 prostate cancer #15673 AR095 Small Intestine Small Intestine (CLONTECH ™) (CLONTECH ™) AR096 Spleen Spleen AR097 Thymus T cells activated Thymus T cells activated AR098 Thymus T cells resting Thymus T cells resting AR099 Tonsil Tonsil AR100 Tonsil geminal center Tonsil geminal center centroblast centroblast AR101 Tonsil germinal center B cell Tonsil germinal center B cell AR102 Tonsil lymph node Tonsil lymph node AR103 Tonsil memory B cell Tonsil memory B cell AR104 Whole Brain Whole Brain AR105 Xenograft ES-2 Xenograft ES-2 AR106 Xenograft SW626 Xenograft SW626 AR119 001: IL-2 001: IL-2 AR120 001: IL-2.1 001: IL-2.1 AR121 001: IL-2_b 001: IL-2_b AR124 002: Monocytes untreated 002: Monocytes untreated (1 hr) (1 hr) AR125 002: Monocytes untreated 002: Monocytes untreated (5 hrs) (5 hrs) AR126 002: Control.1C 002: Control.1C AR127 002: IL2.1C 002: IL2.1C AR130 003: Placebo-treated Rat 003: Placebo-treated Rat Lacrimal Gland Lacrimal Gland AR131 003: Placebo-treated Rat 003: Placebo-treated Rat Submandibular Gland Submandibular Gland AR135 004: Monocytes untreated 004: Monocytes untreated (5 hrs) (5 hrs) AR136 004: Monocytes untreated 004: Monocytes untreated 1 hr 1 hr AR139 005: Placebo (48 hrs) 005: Placebo (48 hrs) AR140 006: pC4 (24 hrs) 006: pC4 (24 hrs) AR141 006: pC4 (48 hrs) 006: pC4 (48 hrs) AR152 007: PHA(1 hr) 007: PHA(1 hr) AR153 007: PHA(6 HRS) 007: PHA(6 HRS) AR154 007: PMA(6 hrs) 007: PMA(6 hrs) AR155 008: 1449_#2 008: 1449_#2 AR161 01: A - max 24 01: A - max 24 AR162 01: A - max 26 01: A - max 26 AR163 01: A - max 30 01: A - max 30 AR164 01: B - max 24 01: B - max 24 AR165 01: B - max 26 01: B - max 26 AR166 01: B - max 30 01: B - max 30 AR167 1449 Sample 1449 Sample AR168 3T3P10 1.0 μM insulin 3T3P10 1.0 μM insulin AR169 3T3P10 10 nM Insulin 3T3P10 10 nM Insulin AR170 3T3P10 10 μM insulin 3T3P10 10 μM insulin AR171 3T3P10 No Insulin 3T3P10 No Insulin AR172 3T3P4 3T3P4 AR173 Adipose (41892) Adipose (41892) AR174 Adipose Diabetic (41611) Adipose Diabetic (41611) AR175 Adipose Diabetic (41661) Adipose Diabetic (41661) AR176 Adipose Diabetic (41689) Adipose Diabetic (41689) AR177 Adipose Diabetic (41706) Adipose Diabetic (41706) AR178 Adipose Diabetic (42352) Adipose Diabetic (42352) AR179 Adipose Diabetic (42366) Adipose Diabetic (42366) AR180 Adipose Diabetic (42452) Adipose Diabetic (42452) AR181 Adipose Diabetic (42491) Adipose Diabetic (42491) AR182 Adipose Normal (41843) Adipose Normal (41843) AR183 Adipose Normal (41893) Adipose Normal (41893) AR184 Adipose Normal (42452) Adipose Normal (42452) AR185 Adrenal Gland Adrenal Gland AR186 Adrenal Gland + Whole Adrenal Gland + Whole Brain Brain AR187 B7(1 hr) + (inverted) B7(1 hr) + (inverted) AR188 Breast (18275A2B) Breast (18275A2B) AR189 Breast (4004199) Breast (4004199) AR190 Breast (4004399) Breast (4004399) AR191 Breast (4004943B7) Breast (4004943B7) AR192 Breast (4005570B1) Breast (4005570B1) AR193 Breast Cancer (4004127A30) Breast Cancer (4004127A30) AR194 Breast Cancer (400443A21) Breast Cancer (400443A21) AR195 Breast Cancer (4004643A2) Breast Cancer (4004643A2) AR196 Breast Cancer (4004710A7) Breast Cancer (4004710A7) AR197 Breast Cancer (4004943A21) Breast Cancer (4004943A21) AR198 Breast Cancer (400553A2) Breast Cancer (400553A2) AR199 Breast Cancer (9805C046R) Breast Cancer (9805C046R) AR200 Breast Cancer (9806C012R) Breast Cancer (9806C012R) AR201 Breast Cancer (ODQ 45913) Breast Cancer (ODQ 45913) AR202 Breast Cancer (ODQ45913) Breast Cancer (ODQ45913) AR203 Breast Cancer (ODQ4591B) Breast Cancer (ODQ4591B) AR204 Colon Cancer (15663) Colon Cancer (15663) AR205 Colon Cancer (4005144A4) Colon Cancer (4005144A4) AR206 Colon Cancer (4005413A4) Colon Cancer (4005413A4) AR207 Colon Cancer (4005570B1) Colon Cancer (4005570B1) AR208 Control RNA #1 Control RNA #1 AR209 Control RNA #2 Control RNA #2 AR210 Cultured Preadipocyte (blue) Cultured Preadipocyte (blue) AR211 Cultured Preadipocyte (Red) Cultured Preadipocyte (Red) AR212 Donor II B-Cells 24 hrs Donor II B-Cells 24 hrs AR213 Donor II Resting B-Cells Donor II Resting B-Cells AR214 H114EP12 10 nM Insulin H114EP12 10 nM Insulin AR215 H114EP12 (10 nM insulin) H114EP12 (10 nM insulin) AR216 H114EP12 (2.6ug/ul) H114EP12 (2.6ug/ul) AR217 H114EP12 (3.6ug/ul) H114EP12 (3.6ug/ul) AR218 HUVEC #1 HUVEC #1 AR219 HUVEC #2 HUVEC #2 AR221 L6 undiff. L6 undiff. AR222 L6 Undifferentiated L6 Undifferentiated AR223 L6P8 + 10 nM Insulin L6P8 + 10 nM Insulin AR224 L6P8 + HS L6P8 + HS AR225 L6P8 10 nM Insulin L6P8 10 nM Insulin AR226 Liver (00-06-A007B) Liver (00-06-A007B) AR227 Liver (96-02-A075) Liver (96-02-A075) AR228 Liver (96-03-A144) Liver (96-03-A144) AR229 Liver (96-04-A138) Liver (96-04-A138) AR230 Liver (97-10-A074B) Liver (97-10-A074B) AR231 Liver (98-09-A242A) Liver (98-09-A242A) AR232 Liver Diabetic (1042) Liver Diabetic (1042) AR233 Liver Diabetic (41616) Liver Diabetic (41616) AR234 Liver Diabetic (41955) Liver Diabetic (41955) AR235 Liver Diabetic (42352R) Liver Diabetic (42352R) AR236 Liver Diabetic (42366) Liver Diabetic (42366) AR237 Liver Diabetic (42483) Liver Diabetic (42483) AR238 Liver Diabetic (42491) Liver Diabetic (42491) AR239 Liver Diabetic (99-09- Liver Diabetic (99-09-A281A) A281A) AR240 Lung Lung AR241 Lung (27270) Lung (27270) AR242 Lung (2727Q) Lung (2727Q) AR243 Lung Cancer (4005116A1) Lung Cancer (4005116A1) AR244 Lung Cancer (4005121A5) Lung Cancer (4005121A5) AR245 Lung Cancer (4005121A5)) Lung Cancer (4005121A5)) AR246 Lung Cancer (4005340A4) Lung Cancer (4005340A4) AR247 Mammary Gland Mammary Gland AR248 Monocyte (CT) Monocyte (CT) AR249 Monocyte (OCT) Monocyte (OCT) AR250 Monocytes (CT) Monocytes (CT) AR251 Monocytes (INFG 18 hr) Monocytes (INFG 18 hr) AR252 Monocytes (INFG 18 hr) Monocytes (INFG 18 hr) AR253 Monocytes (INFG 8-11) Monocytes (INFG 8-11) AR254 Monocytes (O CT) Monocytes (O CT) AR255 Muscle (91-01-A105) Muscle (91-01-A105) AR256 Muscle (92-04-A059) Muscle (92-04-A059) AR257 Muscle (97-11-A056d) Muscle (97-11-A056d) AR258 Muscle (99-06-A210A) Muscle (99-06-A210A) AR259 Muscle (99-07-A203B) Muscle (99-07-A203B) AR260 Muscle (99-7-A203B) Muscle (99-7-A203B) AR261 Muscle Diabetic (42352R) Muscle Diabetic (42352R) AR262 Muscle Diabetic (42366) Muscle Diabetic (42366) AR263 NK-19 Control NK-19 Control AR264 NK-19 IL Treated 72 hrs NK-19 IL Treated 72 hrs AR265 NK-19 UK Treated 72 hrs. NK-19 UK Treated 72 hrs. AR266 Omentum Normal (94-08- Omentum Normal (94-08- B009) B009) AR267 Omentum Normal (97-01- Omentum Normal (97-01- A039A) A039A) AR268 Omentum Normal (97-04- Omentum Normal (97-04- A114C) A114C) AR269 Omentum Normal (97-06- Omentum Normal (97-06- A117C) A117C) AR270 Omentum Normal (97-09- Omentum Normal (97-09- B004C) B004C) AR271 Ovarian Cancer (17717AID) Ovarian Cancer (17717AID) AR272 Ovarian Cancer Ovarian Cancer (9905C023RC) (9905C023RC) AR273 Ovarian Cancer Ovarian Cancer (9905C032RC) (9905C032RC) AR274 Ovary (9508G045) Ovary (9508G045) AR275 Ovary (9701G208) Ovary (9701G208) AR276 Ovary 9806G005 Ovary 9806G005 AR277 Pancreas Pancreas AR278 Placebo Placebo AR279 rIL2 Control rIL2 Control AR280 RSS288L RSS288L AR281 RSS288LC RSS288LC AR282 Salivary Gland Salivary Gland AR283 Skeletal Muscle Skeletal Muscle AR284 Skeletal Muscle (91-01- Skeletal Muscle (91-01-A105) A105) AR285 Skeletal Muscle (42180) Skeletal Muscle (42180) AR286 Skeletal Muscle (42386) Skeletal Muscle (42386) AR287 Skeletal Muscle (42461) Skeletal Muscle (42461) AR288 Skeletal Muscle (91-01- Skeletal Muscle (91-01-A105) A105) AR289 Skeletal Muscle (92-04- Skeletal Muscle (92-04-A059) A059) AR290 Skeletal Muscle (96-08- Skeletal Muscle (96-08-A171) A171) AR291 Skeletal Muscle (97-07- Skeletal Muscle (97-07- A190A) A190A) AR292 Skeletal Muscle Diabetic Skeletal Muscle Diabetic (42352) (42352) AR293 Skeletal Muscle Diabetic Skeletal Muscle Diabetic (42366) (42366) AR294 Skeletal Muscle Diabetic Skeletal Muscle Diabetic (42395) (42395) AR295 Skeletal Muscle Diabetic Skeletal Muscle Diabetic (42483) (42483) AR296 Skeletal Muscle Diabetic Skeletal Muscle Diabetic (42491) (42491) AR297 Skeletal Muscle Diabetic Skeletal Muscle Diabetic 42352 42352 AR298 Skeletal Musle (42461) Skeletal Musle (42461) AR299 Small Intestine Small Intestine AR300 Stomach Stomach AR301 T-Cell + HDPBQ71.fc 1449 T-Cell + HDPBQ71.fc 1449 16 hrs 16 hrs AR302 T-Cell + HDPBQ71.fc 1449 T-Cell + HDPBQ71.fc 1449 6 hrs 6 hrs AR303 T-Cell + IL2 16 hrs T-Cell + IL2 16 hrs AR304 T-Cell + IL2 6 hrs T-Cell + IL2 6 hrs AR306 T-Cell Untreated 16 hrs T-Cell Untreated 16 hrs AR307 T-Cell Untreated 6 hrs T-Cell Untreated 6 hrs AR308 T-Cells 24 hours T-Cells 24 hours AR309 T-Cells 24 hrs T-Cells 24 hrs AR310 T-Cells 24 hrs. T-Cells 24 hrs. AR311 T-Cells 24 hrs T-Cells 24 hrs AR312 T-Cells 4 days T-Cells 4 days AR313 Thymus Thymus AR314 TRE TRE AR315 TREC TREC AR317 B lymphocyte, B lymphocyte, AR318 (non-T; non-B) (non-T; non-B) AR326 001-293 RNA (Vector 001-293 RNA (Vector Control) Control) AR327 001: Control 001: Control AR328 001: Control.1 001: Control.1 AR355 Acute Lymphocyte Leukemia Acute Lymphocyte Leukemia AR356 AML Patient #11 AML Patient #11 AR357 AML Patient #2 AML Patient #2 AR358 AML Patient #2 SGAH AML Patient #2 SGAH AR359 AML Patient#2 AML Patient#2 AR360 Aorta Aorta AR361 B Cell B Cell AR362 B lymphoblast B lymphoblast AR363 B lymphocyte B lymphocyte AR364 B lymphocytes B lymphocytes AR365 B-cell B-cell AR366 B-Cells B-Cells AR367 B-Lymphoblast B-Lymphoblast AR368 B-Lymphocytes B-Lymphocytes AR369 Bladder Bladder AR370 Bone Marrow Bone Marrow AR371 Bronchial Epithelial Cell Bronchial Epithelial Cell AR372 Bronchial Epithelial Cells Bronchial Epithelial Cells AR373 Caco-2A Caco-2A AR374 Caco-2B Caco-2B AR375 Caco-2C Caco-2C AR376 Cardiac #1 Cardiac #1 AR377 Cardiac #2 Cardiac #2 AR378 Chest Muscle Chest Muscle AR381 Dendritic Cell Dendritic Cell AR382 Dendritic cells Dendritic cells AR383 E. coli E. coli AR384 Epithelial Cells Epithelial Cells AR385 Esophagus Esophagus AR386 FPPS FPPS AR387 FPPSC FPPSC AR388 HepG2 Cell Line HepG2 Cell Line AR389 HepG2 Cell line Buffer 1 hr. HepG2 Cell line Buffer 1 hr. AR390 HepG2 Cell line Buffer 06 hr HepG2 Cell line Buffer 06 hr AR391 HepG2 Cell line Buffer 24 hr. HepG2 Cell line Buffer 24 hr. AR392 HepG2 Cell line Insulin 01 hr. HepG2 Cell line Insulin 01 hr. AR393 HepG2 Cell line Insulin 06 hr. HepG2 Cell line Insulin 06 hr. AR394 HepG2 Cell line Insulin 24 hr. HepG2 Cell line Insulin 24 hr. AR398 HMC-1 HMC-1 AR399 HMCS HMCS AR400 HMSC HMSC AR401 HUVEC #3 HUVEC #3 AR402 HUVEC #4 HUVEC #4 AR404 KIDNEY NORMAL KIDNEY NORMAL AR405 KIDNEY TUMOR KIDNEY TUMOR AR406 KIDNEY TUMOR AR407 Lymph Node Lymph Node AR408 Macrophage Macrophage AR409 Megakarioblast Megakarioblast AR410 Monocyte Monocyte AR411 Monocytes Monocytes AR412 Myocardium Myocardium AR413 Myocardium #3 Myocardium #3 AR414 Myocardium #4 Myocardium #4 AR415 Myocardium #5 Myocardium #5 AR416 NK NK AR417 NK cell NK cell AR418 NK cells NK cells AR419 NKYa NKYa AR420 NKYa019 NKYa019 AR421 Ovary Ovary AR422 Patient #11 Patient #11 AR423 Peripheral blood Peripheral blood AR424 Primary Adipocytes Primary Adipocytes AR425 Promyeloblast Promyeloblast AR427 RSSWT RSSWT AR428 RSSWTC RSSWTC AR429 SW 480(G1) SW 480(G1) AR430 SW 480(G2) SW 480(G2) AR431 SW 480(G3) SW 480(G3) AR432 SW 480(G4) SW 480(G4) AR433 SW 480(G5) SW 480(G5) AR434 T Lymphoblast T Lymphoblast AR435 T Lymphocyte T Lymphocyte AR436 T-Cell T-Cell AR438 T-Cell, T-Cell, AR439 T-Cells T-Cells AR440 T-lymphoblast T-lymphoblast AR441 Th 1 Th 1 AR442 Th 2 Th 2 AR443 Th1 Th1 AR444 Th2 Th2 H0002 Human Adult Heart Human Adult Heart H0004 Human Adult Spleen Human Adult Spleen H0008 Whole 6 Week Old Embryo H0009 Human Fetal Brain H0011 Human Fetal Kidney Human Fetal Kidney H0012 Human Fetal Kidney Human Fetal Kidney H0013 Human 8 Week Whole Human 8 Week Old Embryo Embryo H0014 Human Gall Bladder Human Gall Bladder H0015 Human Gall Bladder, fraction Human Gall Bladder II H0016 Human Greater Omentum Human Greater Omentum H0020 Human Hippocampus Human Hippocampus H0022 Jurkat Cells Jurkat T-Cell Line H0023 Human Fetal Lung H0024 Human Fetal Lung III Human Fetal Lung H0026 Namalwa Cells Namalwa B-Cell Line, EBV immortalized H0028 Human Old Ovary Human Old Ovary H0030 Human Placenta H0031 Human Placenta Human Placenta H0032 Human Prostate Human Prostate H0033 Human Pituitary Human Pituitary H0036 Human Adult Small Intestine Human Adult Small Intestine H0038 Human Testes Human Testes H0039 Human Pancreas Tumor Human Pancreas Tumor H0040 Human Testes Tumor Human Testes Tumor H0041 Human Fetal Bone Human Fetal Bone H0042 Human Adult Pulmonary Human Adult Pulmonary H0044 Human Cornea Human Cornea H0045 Human Esophagus, Cancer Human Esophagus, cancer H0046 Human Endometrial Tumor Human Endometrial Tumor H0047 Human Fetal Liver Human Fetal Liver H0048 Human Pineal Gland Human Pineal Gland H0050 Human Fetal Heart Human Fetal Heart H0051 Human Hippocampus Human Hippocampus H0052 Human Cerebellum Human Cerebellum H0056 Human Umbilical Vein, Human Umbilical Vein Endo. remake Endothelial Cells H0057 Human Fetal Spleen H0058 Human Thymus Tumor Human Thymus Tumor H0059 Human Uterine Cancer Human Uterine Cancer H0060 Human Macrophage Human Macrophage H0061 Human Macrophage Human Macrophage H0063 Human Thymus Human Thymus H0065 Human Esophagus, Normal Human Esophagus, normal H0068 Human Skin Tumor Human Skin Tumor H0069 Human Activated T-Cells Activated T-Cells H0070 Human Pancreas Human Pancreas H0071 Human Infant Adrenal Gland Human Infant Adrenal Gland H0073 Human Leiomyeloid Human Leiomyeloid Carcinoma Carcinoma H0075 Human Activated T-Cells (II) Activated T-Cells H0076 Human Membrane Bound Human Membrane Bound Polysomes Polysomes H0078 Human Lung Cancer Human Lung Cancer H0081 Human Fetal Epithelium Human Fetal Skin (Skin) H0083 HUMAN JURKAT Jurkat Cells MEMBRANE BOUND POLYSOMES H0085 Human Colon Human Colon H0086 Human epithelioid sarcoma Epithelioid Sarcoma, muscle H0087 Human Thymus Human Thymus H0090 Human T-Cell Lymphoma T-Cell Lymphoma H0097 Human Adult Heart, Human Adult Heart subtracted H0098 Human Adult Liver, Human Adult Liver subtracted H0099 Human Lung Cancer, Human Lung Cancer subtracted H0100 Human Whole Six Week Old Human Whole Six Week Old Embryo Embryo H0101 Human 7 Weeks Old Human Whole 7 Week Old Embryo, subtracted Embryo H0102 Human Whole 6 Week Old Human Whole Six Week Old Embryo (II), subt Embryo H0103 Human Fetal Brain, Human Fetal Brain subtracted H0105 Human Fetal Heart, Human Fetal Heart subtracted H0107 Human Infant Adrenal Human Infant Adrenal Gland Gland, subtracted H0108 Human Adult Lymph Node, Human Adult Lymph Node subtracted H0109 Human Macrophage, Macrophage subtracted H0110 Human Old Ovary, Human Old Ovary subtracted H0111 Human Placenta, subtracted Human Placenta H0116 Human Thymus Tumor, Human Thymus Tumor subtracted H0118 Human Adult Kidney Human Adult Kidney H0119 Human Pediatric Kidney Human Pediatric Kidney H0120 Human Adult Spleen, Human Adult Spleen subtracted H0121 Human Cornea, subtracted Human Cornea H0122 Human Adult Skeletal Human Skeletal Muscle Muscle H0123 Human Fetal Dura Mater Human Fetal Dura Mater H0124 Human Rhabdomyosarcoma Human Rhabdomyosarcoma H0125 Cem cells cyclohexamide Cyclohexamide Treated Cem, treated Jurkat, Raji, and Supt H0128 Jurkat cells, thiouridine Jurkat Cells activated H0130 LNCAP untreated LNCAP Cell Line H0131 LNCAP + o.3 nM R1881 LNCAP Cell Line H0132 LNCAP + 30 nM R1881 LNCAP Cell Line H0134 Raji Cells, cyclohexamide Cyclohexamide Treated Cem, treated Jurkat, Raji, and Supt H0135 Human Synovial Sarcoma Human Synovial Sarcoma H0136 Supt Cells, cyclohexamide Cyclohexamide Treated Cem, treated Jurkat, Raji, and Supt H0139 Activated T-Cells, 4 hrs. Activated T-Cells H0140 Activated T-Cells, 8 hrs. Activated T-Cells H0141 Activated T-Cells, 12 hrs. Activated T-Cells H0144 Nine Week Old Early Stage 9 Wk Old Early Stage Human Human H0147 Human Adult Liver Human Adult Liver H0149 7 Week Old Early Stage Human Whole 7 Week Old Human, subtracted Embryo H0150 Human Epididymus Epididymis H0151 Early Stage Human Liver Human Fetal Liver H0154 Human Fibrosarcoma Human Skin Fibrosarcoma H0156 Human Adrenal Gland Human Adrenal Gland Tumor Tumor H0158 Activated T-Cells, 4 hrs., Activated T-Cells ligation 2 H0159 Activated T-Cells, 8 hrs., Activated T-Cells ligation 2 H0161 Activated T-Cells, 24 hrs., Activated T-Cells ligation 2 H0163 Human Synovium Human Synovium H0165 Human Prostate Cancer, Human Prostate Cancer, stage Stage B2 B2 H0166 Human Prostate Cancer, Human Prostate Cancer, stage Stage B2 fraction B2 H0167 Activated T-Cells, 24 hrs. Activated T-Cells H0169 Human Prostate Cancer, Human Prostate Cancer, stage C Stage C fraction H0170 12 Week Old Early Stage Twelve Week Old Early Stage Human Human H0171 12 Week Old Early Stage Twelve Week Old Early Stage Human, II Human H0172 Human Fetal Brain, random Human Fetal Brain primed H0176 CAMA1Ee Cell Line CAMA1Ee Cell Line H0177 CAMA1Ee Cell Line CAMA1Ee Cell Line H0178 Human Fetal Brain Human Fetal Brain H0179 Human Neutrophil Human Neutrophil H0180 Human Primary Breast Human Primary Breast Cancer Cancer H0181 Human Primary Breast Human Primary Breast Cancer Cancer H0182 Human Primary Breast Human Primary Breast Cancer Cancer H0183 Human Colon Cancer Human Colon Cancer H0184 Human Colon Cancer, Human Colon Cancer, metasticized to live metasticized to liver H0187 Resting T-Cell T-Cells H0188 Human Normal Breast Human Normal Breast H0189 Human Resting Macrophage Human Macrophage/Monocytes H0190 Human Activated Human Macrophage (LPS) Macrophage/Monocytes H0192 Cem Cells, cyclohexamide Cyclohexamide Treated Cem, treated, subtra Jurkat, Raji, and Supt H0194 Human Cerebellum, Human Cerebellum subtracted H0196 Human Cardiomyopathy, Human Cardiomyopathy subtracted H0197 Human Fetal Liver, Human Fetal Liver subtracted H0199 Human Fetal Liver, Human Fetal Liver subtracted, neg clone H0200 Human Greater Omentum, Human Greater Omentum fract II remake, H0201 Human Hippocampus, Human Hippocampus subtracted H0202 Jurkat Cells, cyclohexamide Cyclohexamide Treated Cem, treated, subtraction Jurkat, Raji, and Supt H0204 Human Colon Cancer, Human Colon Cancer subtracted H0205 Human Colon Cancer, Human Colon Cancer differential H0207 LNCAP, differential LNCAP Cell Line expression H0208 Early Stage Human Lung, Human Fetal Lung subtracted H0209 Human Cerebellum, Human Cerebellum differentially expressed H0211 Human Prostate, differential Human Prostate expression H0212 Human Prostate, subtracted Human Prostate H0213 Human Pituitary, subtracted Human Pituitary H0214 Raji cells, cyclohexamide Cyclohexamide Treated Cem, treated, subtracted Jurkat, Raji, and Supt H0215 Raji cells, cyclohexamide Cyclohexamide Treated Cem, treated, differentially Jurkat, Raji, and Supt expressed H0216 Supt cells, cyclohexamide Cyclohexamide Treated Cem, treated, subtracted Jurkat, Raji, and Supt H0217 Supt cells, cyclohexamide Cyclohexamide Treated Cem, treated, differentially Jurkat, Raji, and Supt expressed H0218 Activated T-Cells, 0 hrs, Activated T-Cells subtracted H0219 Activated T-Cells, 0 hrs, Activated T-Cells differentially expressed H0220 Activated T-Cells, 4 hrs, Activated T-Cells subtracted H0222 Activated T-Cells, 8 hrs, Activated T-Cells subtracted H0223 Activated T-Cells, 8 hrs, Activated T-Cells differentially expressed H0224 Activated T-Cells, 12 hrs, Activated T-Cells subtracted H0225 Activated T-Cells, 12 hrs, Activated T-Cells differentially expressed H0229 Early Stage Human Brain, Early Stage Human Brain random primed H0230 Human Cardiomyopathy, diff Human Cardiomyopathy exp H0231 Human Colon, subtraction Human Colon H0232 Human Colon, differential Human Colon expression H0234 human colon cancer, Human Colon Cancer, metastatic to liver, metasticized to liver differentially expressed H0235 Human colon cancer, Human Colon Cancer, metaticized to liver, metasticized to liver subtraction H0239 Human Kidney Tumor Human Kidney Tumor H0240 C7MCF7 cell line, estrogen C7MCF7 Cell Line, estrogen treated, Differential treated H0241 C7MCF7 cell line, estrogen C7MCF7 Cell Line, estrogen treated, subtraction treated H0242 Human Fetal Heart, Human Fetal Heart Differential (Fetal-Specific) H0244 Human 8 Week Whole Human 8 Week Old Embryo Embryo, subtracted H0247 Human Membrane Bound Human Membrane Bound Polysomes-Enzyme Polysomes Subtraction H0249 HE7, subtracted by Human Whole 7 Week Old hybridization with E7 cDNA Embryo H0250 Human Activated Monocytes Human Monocytes H0251 Human Chondrosarcoma Human Chondrosarcoma H0252 Human Osteosarcoma Human Osteosarcoma H0253 Human adult testis, large Human Adult Testis inserts H0254 Breast Lymph node cDNA Breast Lymph Node library H0255 breast lymph node CDNA Breast Lymph Node library H0256 HL-60, unstimulated Human HL-60 Cells, unstimulated H0257 HL-60, PMA 4H HL-60 Cells, PMA stimulated 4H H0261 H. cerebellum, Enzyme Human Cerebellum subtracted H0263 human colon cancer Human Colon Cancer H0264 human tonsils Human Tonsil H0265 Activated T-Cell T-Cells (12 hs)/Thiouridine labelledEco H0266 Human Microvascular HMEC Endothelial Cells, fract. A H0267 Human Microvascular HMEC Endothelial Cells, fract. B H0268 Human Umbilical Vein HUVE Cells Endothelial Cells, fract. A H0269 Human Umbilical Vein HUVE Cells Endothelial Cells, fract. B H0270 HPAS (human pancreas, Human Pancreas subtracted) H0271 Human Neutrophil, Activated Human Neutrophil—Activated H0272 HUMAN TONSILS, Human Tonsil FRACTION 2 H0274 Human Adult Spleen, Human Adult Spleen fractionII H0275 Human Infant Adrenal Human Infant Adrenal Gland Gland, Subtracted H0280 K562 + PMA (36 hrs) K562 Cell line H0281 Lymph node, abnorm. cell Lymph Node, abnormal cell line (ATCC ™ #7225) line H0282 HBGB''s differential Human Primary Breast Cancer consolidation H0284 Human OB MG63 control Human Osteoblastoma MG63 fraction I cell line H0286 Human OB MG63 treated Human Osteoblastoma MG63 (10 nM E2) fraction I cell line H0288 Human OB HOS control Human Osteoblastoma HOS fraction I cell line H0290 Human OB HOS treated (1 nM Human Osteoblastoma HOS E2) fraction I cell line H0292 Human OB HOS treated (10 nM Human Osteoblastoma HOS E2) fraction I cell line H0293 WI 38 cells H0294 Amniotic Cells - TNF Amniotic Cells - TNF induced induced H0295 Amniotic Cells - Primary Amniotic Cells - Primary Culture Culture H0298 HCBB''s differential CAMA1Ee Cell Line consolidation H0305 CD34 positive cells (Cord CD34 Positive Cells Blood) H0306 CD34 depleted Buffy Coat CD34 Depleted Buffy Coat (Cord Blood) (Cord Blood) H0309 Human Chronic Synovitis Synovium, Chronic Synovitis/ Osteoarthritis H0310 human caudate nucleus Brain H0316 HUMAN STOMACH Human Stomach H0318 HUMAN B CELL Human B Cell Lymphoma LYMPHOMA H0320 Human frontal cortex Human Frontal Cortex H0321 HUMAN SCHWANOMA Schwanoma H0327 human corpus colosum Human Corpus Callosum H0328 human ovarian cancer Ovarian Cancer H0329 Dermatofibrosarcoma Dermatofibrosarcoma Protuberance Protuberans H0331 Hepatocellular Tumor Hepatocellular Tumor H0333 Hemangiopericytoma Hemangiopericytoma H0334 Kidney cancer Kidney Cancer H0339 Duodenum Duodenum H0341 Bone Marrow Cell Line Bone Marrow Cell Line RS4; 11 (RS4; 11) H0343 stomach cancer (human) Stomach Cancer - 5383A (human) H0344 Adipose tissue (human) Adipose - 6825A (human) H0345 SKIN Skin - 4000868H H0346 Brain-medulloblastoma Brain (Medulloblastoma)- 9405C006R H0349 human adult liver cDNA Human Adult Liver library H0350 Human Fetal Liver, mixed 10 Human Fetal Liver, mixed & 14 week 10&14 Week H0351 Glioblastoma Glioblastoma H0352 wilm''s tumor Wilm''s Tumor H0354 Human Leukocytes Human Leukocytes H0355 Human Liver Human Liver, normal Adult H0356 Human Kidney Human Kidney H0357 H. Normalized Fetal Liver, II Human Fetal Liver H0359 KMH2 cell line KMH2 H0360 Hemangiopericytoma Hemangiopericytoma H0361 Human rejected kidney Human Rejected Kidney H0362 HeLa cell line HELA CELL LINE H0366 L428 cell line L428 H0369 H. Atrophic Endometrium Atrophic Endometrium and myometrium H0370 H. Lymph node breast Lymph node with Met. Breast Cancer Cancer H0371 Eosinophils- Eosinophils-Hypereosinophilia Hypereosinophilia patient patient H0372 Human Testes Human Testes H0373 Human Heart Human Adult Heart H0374 Human Brain Human Brain H0375 Human Lung Human Lung H0376 Human Spleen Human Adult Spleen H0379 Human Tongue, frac 1 Human Tongue H0380 Human Tongue, frac 2 Human Tongue H0381 Bone Cancer Bone Cancer H0383 Human Prostate BPH, re- Human Prostate BPH excision H0384 Brain, Kozak Human Brain H0386 Leukocyte and Lung; 4 Human Leukocytes screens H0388 Human Rejected Kidney, 704 Human Rejected Kidney re-excision H0390 Human Amygdala Human Amygdala Depression Depression, re-excision H0391 H. Meniingima, M6 Human Meningima H0392 H. Meningima, M1 Human Meningima H0393 Fetal Liver, subtraction II Human Fetal Liver H0394 A-14 cell line Redd-Sternberg cell H0395 A1-CELL LINE Redd-Sternberg cell H0396 L1 Cell line Redd-Sternberg cell H0399 Human Kidney Cortex, re- Human Kidney Cortex rescue H0400 Human Striatum Depression, Human Brain, Striatum re-rescue Depression H0402 CD34 depleted Buffy Coat CD34 Depleted Buffy Coat (Cord Blood), re-excision (Cord Blood) H0403 H. Umbilical Vein HUVE Cells Endothelial Cells, IL4 induced H0404 H. Umbilical Vein HUVE Cells endothelial cells, uninduced H0405 Human Pituitary, subtracted Human Pituitary VI H0406 H Amygdala Depression, Human Amygdala Depression subtracted H0408 Human kidney Cortex, Human Kidney Cortex subtracted H0409 H. Striatum Depression, Human Brain, Striatum subtracted Depression H0410 H. Male bladder, adult H Male Bladder, Adult H0411 H Female Bladder, Adult Human Female Adult Bladder H0412 Human umbilical vein HUVE Cells endothelial cells, IL-4 induced H0413 Human Umbilical Vein HUVE Cells Endothelial Cells, uninduced H0414 Ovarian Tumor I, OV5232 Ovarian Tumor, OV5232 H0415 H. Ovarian Tumor, II, Ovarian Tumor, OV5232 OV5232 H0416 Human Neutrophils, Human Neutrophil - Activated Activated, re-excision H0417 Human Pituitary, subtracted Human Pituitary VIII H0418 Human Pituitary, subtracted Human Pituitary VII H0419 Bone Cancer, re-excision Bone Cancer H0421 Human Bone Marrow, re- Bone Marrow excision H0422 T-Cell PHA 16 hrs T-Cells H0423 T-Cell PHA 24 hrs T-Cells H0424 Human Pituitary, subt IX Human Pituitary H0427 Human Adipose Human Adipose, left hiplipoma H0428 Human Ovary Human Ovary Tumor H0429 K562 + PMA (36 hrs), re- K562 Cell line excision H0431 H. Kidney Medulla, re- Kidney medulla excision H0433 Human Umbilical Vein HUVE Cells Endothelial cells, frac B, re- excision H0434 Human Brain, striatum, re- Human Brain, Striatum excision H0435 Ovarian Tumor 10-3-95 Ovarian Tumor, OV350721 H0436 Resting T-Cell Library, II T-Cells H0437 H Umbilical Vein HUVE Cells Endothelial Cells, frac A, re- excision H0438 H. Whole Brain #2, re- Human Whole Brain #2 excision H0439 Human Eosinophils Eosinophils H0441 H. Kidney Cortex, subtracted Kidney cortex H0443 H. Adipose, subtracted Human Adipose, left hiplipoma H0444 Spleen metastic melanoma Spleen, Metastic malignant melanoma H0445 Spleen, Chronic lymphocytic Human Spleen, CLL leukemia H0449 CD34+ cell, I CD34 positive cells H0453 H. Kidney Pyramid, Kidney pyramids subtracted H0455 H. Striatum Depression, subt Human Brain, Striatum Depression H0457 Human Eosinophils Human Eosinophils H0458 CD34+ cell, I, frac II CD34 positive cells H0459 CD34+cells, II, FRACTION 2 CD34 positive cells H0462 H. Amygdala Depression, subtracted H0477 Human Tonsil, Lib 3 Human Tonsil H0478 Salivary Gland, Lib 2 Human Salivary Gland H0479 Salivary Gland, Lib 3 Human Salivary Gland H0483 Breast Cancer cell line, MDA Breast Cancer Cell line, MDA 36 36 H0484 Breast Cancer Cell line, Breast Cancer Cell line, angiogenic Angiogenic, 36T3 H0485 Hodgkin''s Lymphoma I Hodgkin''s Lymphoma I H0486 Hodgkin''s Lymphoma II Hodgkin''s Lymphoma II H0487 Human Tonsils, lib I Human Tonsils H0488 Human Tonsils, Lib 2 Human Tonsils H0489 Crohn''s Disease Ileum H0490 H1-60, untreated, subtracted Human HL-60 Cells, unstimulated H0491 HL-60, PMA 4H, subtracted HL-60 Cells, PMA stimulated 4H H0492 HL-60, RA 4h, Subtracted HL-60 Cells, RA stimulated for 4H H0494 Keratinocyte Keratinocyte H0497 HEL cell line HEL cell line H0505 Human Astrocyte Human Astrocyte H0506 Ulcerative Colitis Colon H0509 Liver, Hepatoma Human Liver, Hepatoma, patient 8 H0510 Human Liver, normal Human Liver, normal, Patient # 8 H0512 Keratinocyte, lib 3 Keratinocyte H0518 pBMC stimulated w/ poly I/C pBMC stimulated with poly I/C H0519 NTERA2, control NTERA2, Teratocarcinoma cell line H0520 NTERA2 + retinoic acid, 14 NTERA2, Teratocarcinoma cell days line H0521 Primary Dendritic Cells, lib 1 Primary Dendritic cells H0522 Primary Dendritic cells, frac 2 Primary Dendritic cells H0525 PCR, pBMC I/C treated pBMC stimulated with poly I/C H0528 Poly[I]/Poly[C] Normal Lung Poly[I]/Poly[C] Normal Lung Fibroblasts Fibroblasts H0529 Myoloid Progenitor Cell Line TF-1 Cell Line; Myoloid progenitor cell line H0530 Human Dermal Endothelial Human Dermal Endothelial Cells, untreated Cells; untreated H0538 Merkel Cells Merkel cells H0539 Pancreas Islet Cell Tumor Pancreas Islet Cell Tumour H0540 Skin, burned Skin, leg burned H0542 T Cell helper I Helper T cell H0543 T cell helper II Helper T cell H0544 Human endometrial stromal Human endometrial stromal cells cells H0545 Human endometrial stromal Human endometrial stromal cells-treated with cells-treated with proge progesterone H0546 Human endometrial stromal Human endometrial stromal cells-treated with estradiol cells-treated with estra H0547 NTERA2 teratocarcinoma NTERA2, Teratocarcinoma cell cell line + retinoic acid (14 line days) H0549 H. Epididiymus, caput & Human Epididiymus, caput and corpus corpus H0550 H. Epididiymus, cauda Human Epididiymus, cauda H0551 Human Thymus Stromal Human Thymus Stromal Cells Cells H0553 Human Placenta Human Placenta H0555 Rejected Kidney, lib 4 Human Rejected Kidney H0556 Activated T- T-Cells cell(12 h)/Thiouridine-re- excision H0559 HL-60, PMA 4H, re-excision HL-60 Cells, PMA stimulated 4H H0560 KMH2 KMH2 H0561 L428 L428 H0562 Human Fetal Brain, Human Fetal Brain normalized c5-11-26 H0563 Human Fetal Brain, Human Fetal Brain normalized 50021F H0564 Human Fetal Brain, Human Fetal Brain normalized C5001F H0566 Human Fetal Human Fetal Brain Brain, normalized c50F H0567 Human Fetal Brain, Human Fetal Brain normalized A5002F H0569 Human Fetal Brain, Human Fetal Brain normalized CO H0570 Human Fetal Brain, Human Fetal Brain normalized C500H H0571 Human Fetal Brain, Human Fetal Brain normalized C500HE H0572 Human Fetal Brain, Human Fetal Brain normalized AC5002 H0574 Hepatocellular Tumor; re- Hepatocellular Tumor excision H0575 Human Adult Pulmonary; re- Human Adult Pulmonary excision H0576 Resting T-Cell; re-excision T-Cells H0579 Pericardium Pericardium H0580 Dendritic cells, pooled Pooled dendritic cells H0581 Human Bone Marrow, Human Bone Marrow treated H0583 B Cell lymphoma B Cell Lymphoma H0584 Activated T-cells, 24 hrs, re- Activated T-Cells excision H0585 Activated T-Cells, 12 hrs, re- Activated T-Cells excision H0586 Healing groin wound, 6.5 healing groin wound, 6.5 hours hours post incision post incision - 2/ H0587 Healing groin wound; 7.5 Groin-Feb. 19, 1997 hours post incision H0589 CD34 positive cells (cord CD34 Positive Cells blood), re-ex H0590 Human adult small Human Adult Small Intestine intestine, re-excision H0591 Human T-cell lymphoma; re- T-Cell Lymphoma excision H0592 Healing groin wound - zero HGS wound healing project; hr post-incision (control) abdomen H0593 Olfactory Olfactory epithelium from roof epithelium; nasalcavity of left nasal cacit H0594 Human Lung Cancer; re- Human Lung Cancer excision H0595 Stomach cancer (human); re- Stomach Cancer - 5383A excision (human) H0596 Human Colon Cancer; re- Human Colon Cancer excision H0597 Human Colon; re-excision Human Colon H0598 Human Stomach; re-excision Human Stomach H0599 Human Adult Heart; re- Human Adult Heart excision H0600 Healing Abdomen Abdomen wound; 70&90 min post incision H0601 Healing Abdomen Wound; 15 Abdomen days post incision H0602 Healing Abdomen Abdomen Wound; 21&29 days post incision H0604 Human Pituitary, re-excision Human Pituitary H0606 Human Primary Breast Human Primary Breast Cancer Cancer; re-excision H0607 H. Leukocytes, normalized H. Leukocytes cot 50A3 H0609 H. Leukocytes, normalized H. Leukocytes cot >500A H0610 H. Leukocytes, normalized H. Leukocytes cot 5A H0611 H. Leukocytes, normalized H. Leukocytes cot 500 B H0613 H. Leukocytes, normalized H. Leukocytes cot 5B H0614 H. Leukocytes, normalized H. Leukocytes cot 500 A H0615 Human Ovarian Cancer Ovarian Cancer Reexcision H0616 Human Testes, Reexcision Human Testes H0617 Human Primary Breast Human Primary Breast Cancer Cancer Reexcision H0618 Human Adult Testes, Large Human Adult Testis Inserts, Reexcision H0619 Fetal Heart Human Fetal Heart H0620 Human Fetal Kidney; Human Fetal Kidney Reexcision H0622 Human Pancreas Tumor; Human Pancreas Tumor Reexcision H0623 Human Umbilical Vein; Human Umbilical Vein Reexcision Endothelial Cells H0624 12 Week Early Stage Human Twelve Week Old Early Stage II; Reexcision Human H0625 Ku 812F Basophils Line Ku 812F Basophils H0626 Saos2 Cells; Untreated Saos2 Cell Line; Untreated H0627 Saos2 Cells; Vitamin D3 Saos2 Cell Line; Vitamin D3 Treated Treated H0628 Human Pre-Differentiated Human Pre-Differentiated Adipocytes Adipocytes H0629 Human Leukocyte, control Human Normalized leukocyte #2 H0631 Saos2, Dexamethosome Saos2 Cell Line; Treated Dexamethosome Treated H0632 Hepatocellular Tumor; re- Hepatocellular Tumor excision H0633 Lung Carcinoma A549 TNFalpha activated A549 - TNFalpha activated Lung Carcinoma H0634 Human Testes Tumor, re- Human Testes Tumor excision H0635 Human Activated T-Cells, re- Activated T-Cells excision H0637 Dendritic Cells From CD34 Dentritic cells from CD34 cells Cells H0638 CD40 activated monocyte CD40 activated monocyte dendridic cells dendridic cells H0640 Ficolled Human Stromal Ficolled Human Stromal Cells, Cells, Untreated Untreated H0641 LPS activated derived LPS activated monocyte dendritic cells derived dendritic cells H0642 Hep G2 Cells, lambda library Hep G2 Cells H0643 Hep G2 Cells, PCR library Hep G2 Cells H0644 Human Placenta (re-excision) Human Placenta H0645 Fetal Heart, re-excision Human Fetal Heart H0646 Lung, Cancer (4005313 A3): Metastatic squamous cell lung Invasive Poorly carcinoma, poorly di Differentiated Lung Adenocarcinoma, H0647 Lung, Cancer (4005163 B7): Invasive poorly differentiated Invasive, Poorly Diff. lung adenocarcinoma Adenocarcinoma, Metastatic H0648 Ovary, Cancer: (4004562 B6) Papillary Cstic neoplasm of low Papillary Serous Cystic malignant potentia Neoplasm, Low Malignant Pot H0649 Lung, Normal: (4005313 B1) Normal Lung H0650 B-Cells B-Cells H0651 Ovary, Normal: Normal Ovary (9805C040R) H0652 Lung, Normal: (4005313 B1) Normal Lung H0653 Stromal Cells Stromal Cells H0654 Lung, Cancer: (4005313 A3) Metastatic Squamous cell lung Invasive Poorly- Carcinoma poorly dif differentiated Metastatic lung adenoc H0656 B-cells (unstimulated) B-cells (unstimulated) H0657 B-cells (stimulated) B-cells (stimulated) H0658 Ovary, Cancer (9809C332): 9809C332 - Poorly differentiate Poorly differentiated adenocarcinoma H0659 Ovary, Cancer (15395A1F): Grade II Papillary Carcinoma, Grade II Papillary Carcinoma Ovary H0660 Ovary, Cancer: (15799A1F) Poorly differentiated Poorly differentiated carcinoma, ovary carcinoma H0661 Breast, Cancer: (4004943 Breast cancer A5) H0662 Breast, Normal: Normal Breast - #4005522(B2) (4005522B2) H0663 Breast, Cancer: (4005522 Breast Cancer - #4005522(A2) A2) H0664 Breast, Cancer: (9806C012R) Breast Cancer H0665 Stromal cells 3.88 Stromal cells 3.88 H0666 Ovary, Cancer: (4004332 Ovarian Cancer, Sample A2) #4004332A2 H0667 Stromal cells(HBM3.18) Stromal cell(HBM 3.18) H0668 stromal cell clone 2.5 stromal cell clone 2.5 H0670 Ovary, Cancer(4004650 A3): Ovarian Cancer - 4004650A3 Well-Differentiated Micropapillary Serous Carcinoma H0671 Breast, Cancer: Breast Cancer - Sample # (9802C02OE) 9802C02OE H0672 Ovary, Cancer: (4004576 Ovarian Cancer(4004576A8) A8) H0673 Human Prostate Cancer, Human Prostate Cancer, stage Stage B2; re-excision B2 H0674 Human Prostate Cancer, Human Prostate Cancer, stage C Stage C; re-excission H0675 Colon, Cancer: Colon Cancer 9808C064R (9808C064R) H0676 Colon, Cancer: Colon Cancer 9808C064R (9808C064R)-total RNA H0677 TNFR degenerate oligo B-Cells H0678 screened clones from Placenta placental library H0682 Serous Papillary serous papillary Adenocarcinoma adenocarcinoma (9606G304SPA3B) H0683 Ovarian Serous Papillary Serous papillary Adenocarcinoma adenocarcinoma, stage 3C (9804G01 H0684 Serous Papillary Ovarian Cancer-9810G606 Adenocarcinoma H0685 Adenocarcinoma of Ovary, Adenocarcinoma of Ovary, Human Cell Line, # Human Cell Line, # OVCAR- OVCAR-3 H0686 Adenocarcinoma of Ovary, Adenocarcinoma of Ovary, Human Cell Line Human Cell Line, # SW-626 H0687 Human normal Human normal ovary(#9610G215) ovary(#9610G215) H0688 Human Ovarian Human Ovarian Cancer(#9807G017) cancer(#9807G017), mRNA from Maura Ru H0689 Ovarian Cancer Ovarian Cancer, #9806G019 H0690 Ovarian Cancer, #9702G001 Ovarian Cancer, #9702G001 H0691 Normal Ovary, #9710G208 normal ovary, #9710G208 H0693 Normal Prostate Normal Prostate Tissue # #ODQ3958EN ODQ3958EN H0694 Prostate gland Prostate gland, adenocarcinoma adenocarcinoma, mod/diff, gleason H0695 mononucleocytes from mononucleocytes from patient patient at Shady Grove Hospit N0003 Human Fetal Brain Human Fetal Brain N0006 Human Fetal Brain Human Fetal Brain N0007 Human Hippocampus Human Hippocampus N0009 Human Hippocampus, Human Hippocampus prescreened S0001 Brain frontal cortex Brain frontal cortex S0002 Monocyte activated Monocyte-activated S0003 Human Osteoclastoma Osteoclastoma S0004 Prostate Prostate BPH S0006 Neuroblastoma Human Neural Blastoma S0007 Early Stage Human Brain Human Fetal Brain S0010 Human Amygdala Amygdala S0011 STROMAL- Osteoclastoma OSTEOCLASTOMA S0013 Prostate Prostate S0014 Kidney Cortex Kidney cortex S0015 Kidney medulla Kidney medulla S0016 Kidney Pyramids Kidney pyramids S0021 Whole brain Whole brain S0022 Human Osteoclastoma Osteoclastoma Stromal Cells Stromal Cells - unamplified S0023 Human Kidney Cortex - Human Kidney Cortex unamplified S0024 Human Kidney Medulla - Human Kidney Medulla unamplified S0026 Stromal cell TF274 stromal cell S0027 Smooth muscle, serum Smooth muscle treated S0028 Smooth muscle, control Smooth muscle S0029 brain stem Brain stem S0030 Brain pons Brain Pons S0031 Spinal cord Spinal cord S0032 Smooth muscle-ILb induced Smooth muscle S0035 Brain medulla oblongata Brain medulla oblongata S0036 Human Substantia Nigra Human Substantia Nigra S0037 Smooth muscle, IL 1b Smooth muscle induced S0038 Human Whole Brain #2 - Human Whole Brain #2 Oligo dT > 1.5 Kb S0039 Hypothalamus Hypothalamus S0040 Adipocytes Human Adipocytes from Osteoclastoma S0042 Testes Human Testes S0044 Prostate BPH prostate BPH S0045 Endothelial cells-control Endothelial cell S0046 Endothelial-induced Endothelial cell S0048 Human Hypothalamus, Human Hypothalamus, Alzheimer''s Alzheimer''s S0049 Human Brain, Striatum Human Brain, Striatum S0050 Human Frontal Cortex, Human Frontal Cortex, Schizophrenia Schizophrenia S0051 Human Human Hypothalamus, Hypothalmus, Schizophrenia Schizophrenia S0052 neutrophils control human neutrophils S0053 Neutrophils IL-1 and LPS human neutrophil induced induced S0106 STRIATUM DEPRESSION S0110 Brain Amygdala Depression S0112 Hypothalamus S0114 Anergic T-cell Anergic T-cell S0116 Bone marrow Bone marrow S0122 Osteoclastoma-normalized A Osteoclastoma S0124 Smooth muscle-edited A Smooth muscle S0126 Osteoblasts Osteoblasts S0132 Epithelial-TNFa and INF Airway Epithelial induced S0134 Apoptotic T-cell apoptotic cells S0136 PERM TF274 stromal cell S0140 eosinophil-IL5 induced eosinophil S0142 Macrophage-oxLDL macrophage-oxidized LDL treated S0144 Macrophage (GM-CSF Macrophage (GM-CSF treated) treated) S0146 prostate-edited prostate BPH S0148 Normal Prostate Prostate S0150 LNCAP prostate cell line LNCAP Cell Line S0152 PC3 Prostate cell line PC3 prostate cell line S0168 Prostate/LNCAP, subtraction I PC3 prostate cell line S0176 Prostate, normal, subtraction I Prostate S0180 Bone Marrow Stroma, Bone Marrow Stroma, TNF & TNF&LPS ind LPS induced S0182 Human B Cell 8866 Human B-Cell 8866 S0188 Prostate, BPH, Lib 2 Human Prostate BPH S0190 Prostate BPH, Lib 2, Human Prostate BPH subtracted S0192 Synovial Fibroblasts Synovial Fibroblasts (control) S0194 Synovial hypoxia Synovial Fibroblasts S0196 Synovial IL-1/TNF Synovial Fibroblasts stimulated S0206 Smooth Muscle-HASTE Smooth muscle normalized S0208 Messangial cell, frac 1 Messangial cell S0210 Messangial cell, frac 2 Messangial cell S0212 Bone Marrow Stromal Cell, Bone Marrow Stromal untreated Cell, untreated S0214 Human Osteoclastoma, re- Osteoclastoma excision S0216 Neutrophils IL-1 and LPS human neutrophil induced induced S0218 Apoptotic T-cell, re-excision apoptotic cells S0220 H. hypothalamus, frac A; re- Hypothalamus excision S0222 H. Frontal cortex, epileptic; re- H. Brain, Frontal Cortex, excision Epileptic S0242 Synovial Fibroblasts Synovial Fibroblasts (Il1/TNF), subt S0250 Human Osteoblasts II Human Osteoblasts S0260 Spinal Cord, re-excision Spinal cord S0276 Synovial hypoxia-RSF Synovial fobroblasts subtracted (rheumatoid) S0278 H Macrophage (GM-CSF Macrophage (GM-CSF treated) treated), re-excision S0280 Human Adipose Tissue, re- Human Adipose Tissue excision S0282 Brain Frontal Cortex, re- Brain frontal cortex excision S0292 Osteoarthritis (OA-4) Human Osteoarthritic Cartilage S0294 Larynx tumor Larynx tumor S0298 Bone marrow stroma, treated Bone marrow stroma, treatedSB S0300 Frontal lobe, dementia; re- Frontal Lobe excision dementia/Alzheimer''s S0306 Larynx normal #10 261-273 Larynx normal S0308 Spleen/normal Spleen normal S0310 Normal trachea Normal trachea S0312 Human osteoarthritic; fraction Human osteoarthritic cartilage II S0314 Human osteoarthritis; fraction I Human osteoarthritic cartilage S0316 Human Normal Human Normal Cartilage Cartilage, Fraction I S0318 Human Normal Cartilage Human Normal Cartilage Fraction II S0328 Palate carcinoma Palate carcinoma S0330 Palate normal Palate normal S0332 Pharynx carcinoma Pharynx carcinoma S0334 Human Normal Cartilage Human Normal Cartilage Fraction III S0336 Human Normal Cartilage Human Normal Cartilage Fraction IV S0338 Human Osteoarthritic Human osteoarthritic cartilage Cartilage Fraction III S0340 Human Osteoarthritic Human osteoarthritic cartilage Cartilage Fraction IV S0342 Adipocytes; re-excision Human Adipocytes from Osteoclastoma S0344 Macrophage-oxLDL; re- macrophage-oxidized LDL excision treated S0346 Human Amygdala; re- Amygdala excision S0348 Cheek Carcinoma Cheek Carcinoma S0350 Pharynx Carcinoma Pharynx carcinoma S0352 Larynx Carcinoma Larynx carcinoma S0354 Colon Normal II Colon Normal S0356 Colon Carcinoma Colon Carcinoma S0358 Colon Normal III Colon Normal S0360 Colon Tumor II Colon Tumor S0362 Human Gastrocnemius Gastrocnemius muscle S0364 Human Quadriceps Quadriceps muscle S0366 Human Soleus Soleus Muscle S0368 Human Pancreatic Islets of Langerhans Langerhans S0370 Larynx carcinoma II Larynx carcinoma S0372 Larynx carcinoma III Larynx carcinoma S0374 Normal colon Normal colon S0376 Colon Tumor Colon Tumor S0378 Pancreas normal PCA4 No Pancreas Normal PCA4 No S0380 Pancreas Tumor PCA4 Tu Pancreas Tumor PCA4 Tu S0382 Larynx carcinoma IV Larynx carcinoma S0384 Tongue carcinoma Tongue carcinoma S0386 Human Whole Brain, re- Whole brain excision S0388 Human Human Hypothalamus, Hypothalamus, schizophrenia, Schizophrenia re-excision S0390 Smooth muscle, control; re- Smooth muscle excision S0392 Salivary Gland Salivary gland; normal S0394 Stomach; normal Stomach; normal S0398 Testis; normal Testis; normal S0400 Brain; normal Brain; normal S0402 Adrenal Gland, normal Adrenal gland; normal S0404 Rectum normal Rectum, normal S0406 Rectum tumour Rectum tumour S0408 Colon, normal Colon, normal S0410 Colon, tumour Colon, tumour S0412 Temporal cortex-Alzheizmer; Temporal cortex, alzheimer subtracted S0414 Hippocampus, Alzheimer Hippocampus, Alzheimer Subtracted Subtracted S0418 CHME Cell Line; treated 5 hrs CHME Cell Line; treated S0420 CHME Cell Line, untreated CHME Cell line, untreatetd S0422 Mo7e Cell Line GM-CSF Mo7e Cell Line GM-CSF treated (1 ng/ml) treated (1 ng/ml) S0424 TF-1 Cell Line GM-CSF TF-1 Cell Line GM-CSF Treated Treated S0426 Monocyte activated; re- Monocyte-activated excision S0428 Neutrophils control; re- human neutrophils excision S0430 Aryepiglottis Normal Aryepiglottis Normal S0432 Sinus piniformis Tumour Sinus piniformis Tumour S0434 Stomach Normal Stomach Normal S0436 Stomach Tumour Stomach Tumour S0438 Liver Normal Met5No Liver Normal Met5No S0440 Liver Tumour Met 5 Tu Liver Tumour S0442 Colon Normal Colon Normal S0444 Colon Tumor Colon Tumour S0446 Tongue Tumour Tongue Tumour S0448 Larynx Normal Larynx Normal S0450 Larynx Tumour Larynx Tumour S0452 Thymus Thymus S0454 Placenta Placenta S0456 Tongue Normal Tongue Normal S0458 Thyroid Normal (SDCA2 Thyroid normal No) S0460 Thyroid Tumour Thyroid Tumour S0462 Thyroid Thyroiditis Thyroid Thyroiditis S0464 Larynx Normal Larynx Normal S0466 Larynx Tumor Larynx Tumor S0468 Ea.hy.926 cell line Ea.hy.926 cell line S0470 Adenocarcinoma PYFD S0472 Lung Mesothelium PYBT S0474 Human blood platelets Platelets S0665 Human Amygdala; re- Amygdala excission S3012 Smooth Muscle Serum Smooth muscle Treated, Norm S3014 Smooth muscle, serum Smooth muscle induced, re-exc S6014 H. hypothalamus, frac A Hypothalamus S6016 H. Frontal Cortex, Epileptic H. Brain, Frontal Cortex, Epileptic S6022 H. Adipose Tissue Human Adipose Tissue S6024 Alzheimers, spongy change Alzheimer''s/Spongy change S6026 Frontal Lobe, Dementia Frontal Lobe dementia/Alzheimer''s S6028 Human Manic Depression Human Manic depression tissue Tissue T0002 Activated T-cells Activated T-Cell, PBL fraction T0003 Human Fetal Lung Human Fetal Lung T0004 Human White Fat Human White Fat T0006 Human Pineal Gland Human Pinneal Gland T0008 Colorectal Tumor Colorectal Tumor T0010 Human Infant Brain Human Infant Brain T0023 Human Pancreatic Human Pancreatic Carcinoma Carcinoma T0039 HSA 172 Cells Human HSA172 cell line T0040 HSC172 cells SA172 Cells T0041 Jurkat T-cell G1 phase Jurkat T-cell T0042 Jurkat T-Cell, S phase Jurkat T-Cell Line T0048 Human Aortic Endothelium Human Aortic Endothilium T0049 Aorta endothelial cells + Aorta endothelial cells TNF-a T0060 Human White Adipose Human White Fat T0067 Human Thyroid Human Thyroid T0068 Normal Ovary, Normal Ovary, Premenopausal Premenopausal T0069 Human Uterus, normal Human Uterus, normal T0071 Human Bone Marrow Human Bone Marrow T0079 Human Kidney, normal Human Kidney, normal Adult Adult T0082 Human Adult Retina Human Adult Retina T0086 Human Pancreatic Human Pancreatic Carcinoma Carcinoma -- Screened T0103 Human colon carcinoma (HCC) cell line T0104 HCC cell line metastisis to liver T0109 Human (HCC) cell line liver (mouse) metastasis, remake T0110 Human colon carcinoma (HCC) cell line, remake T0112 Human (Caco-2) cell line, adenocarcinoma, colon T0114 Human (Caco-2) cell line, adenocarcinoma, colon, remake T0115 Human Colon Carcinoma (HCC) cell line L0002 Atrium cDNA library Human heart L0005 CLONTECH ™ human aorta polyA+ mRNA (#6572) L0015 Human L0021 Human adult (K. Okubo) L0022 Human adult lung 3″ directed MboI cDNA L0024 Human brain ARSanders L0040 Human colon mucosa L0041 Human epidermal keratinocyte L0045 Human keratinocyte differential display (B. Lin) L0053 Human pancreatic tumor L0055 Human promyelocyte L0065 Liver HepG2 cell line. L0070 Selected chromosome 21 cDNA library L0096 Subtracted human retina L0097 Subtracted human retinal pigment epithelium (RPE) L0103 DKFZphamyl amygdala L0105 Human aorta polyA+ aorta (TFujiwara) L0142 Human placenta cDNA placenta (TFujiwara) L0143 Human placenta polyA+ placenta (TFujiwara) L0151 Human testis (C. De Smet) testis L0157 Human fetal brain (TFujiwara) L0163 Human heart cDNA (YNakamura) L0182 Human HeLa (Y. Wang) L0187 Human fibrosarcoma cell line fibrosarcoma HT1080 L0194 Human pancreatic cancer cell pancreatic cancer line Patu 8988t L0295 Human liver EST (Y. L. Yu) L0309 Human E8CASS breast adenocarcinoma L0351 Infant brain, Bento Soares L0352 Normalized infant brain, Bento Soares L0355 P, Human foetal Brain Whole tissue L0356 S, Human foetal Adrenals tissue L0361 STRATAGENE ™ ovary (#937217) L0362 STRATAGENE ™ ovarian cancer (#937219) L0363 NCI_CGAP_GC2 germ cell tumor L0364 NCI_CGAP_GC5 germ cell tumor L0365 NCI_CGAP_Phe1 pheochromocytoma L0366 STRATAGENE ™ schizo schizophrenic brain S-11 brain S11 frontal lobe L0367 NCI_CGAP_Sch1 Schwannoma tumor L0368 NCI_CGAP_SS1 synovial sarcoma L0369 NCI_CGAP_AA1 adrenal adenoma L0370 Johnston frontal cortex pooled frontal lobe L0371 NCI_CGAP_Br3 breast tumor L0372 NCI_CGAP_Co12 colon tumor L0373 NCI_CGAP_Co11 tumor L0374 NCI_CGAP_Co2 tumor L0375 NCI_CGAP_Kid6 kidney tumor L0376 NCI_CGAP_Lar1 larynx L0378 NCI_CGAP_Lu1 lung tumor L0379 NCI_CGAP_Lym3 lymphoma L0381 NCI_CGAP_HN4 squamous cell carcinoma L0382 NCI_CGAP_Pr25 epithelium (cell line) L0383 NCI_CGAP_Pr24 invasive tumor (cell line) L0384 NCI_CGAP_Pr23 prostate tumor L0385 NCI_CGAP_Gas1 gastric tumor L0386 NCI_CGAP_HN3 squamous cell carcinoma from base of tongue L0387 NCI_CGAP_GCB0 germinal center B-cells L0388 NCI_CGAP_HN6 normal gingiva (cell line from immortalized kerati L0389 NCI_CGAP_HN5 normal gingiva (cell line from primary keratinocyt L0394 H, Human adult Brain Cortex tissue L0404 b4HB3MA Cot109 + 103 + 85- Bio L0411 1-NIB L0415 b4HB3MA Cot8-HAP-Ft L0418 b4HB3MA-Cot109 + 10-Bio L0428 Cot1374Ft-4HB3MA L0434 Infant brain library of Dr. M. Soares L0435 Infant brain, LLNL array of Dr. M. Soares 1NIB L0438 normalized infant brain total brain cDNA L0439 Soares infant brain 1NIB L0443 b4HB3MK L0446 N4HB3MK L0454 CLONTECH ™ adult human fat cell library HL1108A L0455 Human retina cDNA retina randomly primed sublibrary L0456 Human retina cDNA retina Tsp509I-cleaved sublibrary L0457 multi-tissue normalized multi-tissue short-fragment L0459 Adult heart, CLONTECH ™ L0460 Adult heart, Lambda gt11 L0462 WATM1 L0463 fetal brain cDNA brain L0465 TEST1, Human adult Testis tissue L0468 HE6W L0471 Human fetal heart, Lambda ZAP Express L0475 KG1-a Lambda Zap Express cDNA library L0476 Fetal brain, STRATAGENE ™ L0480 STRATAGENE ™ cat#937212 (1992) L0481 CD34+DIRECTIONAL L0483 Human pancreatic islet L0485 STRATAGENE ™ Human skeletal muscle skeletal muscle cDNA library, cat. #936215. L0492 Human Genomic L0493 NCI_CGAP_Ov26 papillary serous carcinoma L0497 NCI_CGAP_HSC4 CD34+, CD38− from normal bone marrow donor L0498 NCI_CGAP_HSC3 CD34+, T negative, patient with chronic myelogenou L0499 NCI_CGAP_HSC2 stem cell 34+/38+ L0500 NCI_CGAP_Brn20 oligodendroglioma L0502 NCI_CGAP_Br15 adenocarcinoma L0503 NCI_CGAP_Br17 adenocarcinoma L0504 NCI_CGAP_Br13 breast carcinoma in situ L0505 NCI_CGAP_Br12 invasive carcinoma L0506 NCI_CGAP_Br16 lobullar carcinoma in situ L0507 NCI_CGAP_Br14 normal epithelium L0508 NCI_CGAP_Lu25 bronchioalveolar carcinoma L0509 NCI_CGAP_Lu26 invasive adenocarcinoma L0511 NCI_CGAP_Ov34 borderline ovarian carcinoma L0512 NCI_CGAP_Ov36 borderline ovarian carcinoma L0513 NCI_CGAP_Ov37 early stage papillary serous carcinoma L0514 NCI_CGAP_Ov31 papillary serous carcinoma L0515 NCI_CGAP_Ov32 papillary serous carcinoma L0517 NCI_CGAP_Pr1 L0518 NCI_CGAP_Pr2 L0519 NCI_CGAP_Pr3 L0520 NCI_CGAP_Alv1 alveolar rhabdomyosarcoma L0521 NCI_CGAP_Ew1 Ewing''s sarcoma L0522 NCI_CGAP_Kid1 kidney L0523 NCI_CGAP_Lip2 liposarcoma L0524 NCI_CGAP_Li1 liver L0525 NCI_CGAP_Li2 liver L0526 NCI_CGAP_Pr12 metastatic prostate bone lesion L0527 NCI_CGAP_Ov2 ovary L0528 NCI_CGAP_Pr5 prostate L0529 NCI_CGAP_Pr6 prostate L0530 NCI_CGAP_Pr8 prostate L0532 NCI_CGAP_Thy1 thyroid L0533 NCI_CGAP_HSC1 stem cells L0534 Chromosome 7 Fetal Brain brain cDNA Library L0536 NCI_CGAP_Br4 normal ductal tissue L0539 Chromosome 7 Placental cDNA Library L0540 NCI_CGAP_Pr10 invasive prostate tumor L0542 NCI_CGAP_Pr11 normal prostatic epithelial cells L0543 NCI_CGAP_Pr9 normal prostatic epithelial cells L0544 NCI_CGAP_Pr4 prostatic intraepithelial neoplasia - high grade L0545 NCI_CGAP_Pr4.1 prostatic intraepithelial neoplasia - high grade L0546 NCI_CGAP_Pr18 stroma L0547 NCI_CGAP_Pr16 tumor L0549 NCI_CGAP_HN10 carcinoma in situ from retromolar trigone L0550 NCI_CGAP_HN9 normal squamous epithelium from retromolar trigone L0551 NCI_CGAP_HN7 normal squamous epithelium, floor of mouth L0554 NCI_CGAP_Li8 L0558 NCI_CGAP_Ov40 endometrioid ovarian metastasis L0559 NCI_CGAP_Ov39 papillary serous ovarian metastasis L0560 NCI_CGAP_HN12 moderate to poorly differentiated invasive carcino L0561 NCI_CGAP_HN11 normal squamous epithelium L0562 Chromosome 7 HeLa cDNA Library L0564 Jia bone marrow stroma bone marrow stroma L0565 Normal Human Trabecular Bone Bone Cells L0581 STRATAGENE ™ liver (#937224) L0584 STRATAGENE ™ cDNA library Human heart, cat#936208 L0586 HTCDL1 L0587 STRATAGENE ™ colon HT29 (#937221) L0588 STRATAGENE ™ endothelial cell 937223 L0589 STRATAGENE ™ fetal retina 937202 L0590 STRATAGENE ™ fibroblast (#937212) L0591 STRATAGENE ™ HeLa cell s3 937216 L0592 STRATAGENE ™ hNT neuron (#937233) L0593 STRATAGENE ™ neuroepithelium (#937231) L0594 STRATAGENE ™ neuroepithelium NT2RAMI 937234 L0595 STRATAGENE ™ NT2 neuroepithelial cells neuronal precursor 937230 L0596 STRATAGENE ™ colon (#937204) L0597 STRATAGENE ™ corneal stroma (#937222) L0598 Morton Fetal Cochlea cochlea L0599 STRATAGENE ™ lung (#937210) L0600 Weizmann Olfactory olfactory epithelium Epithelium L0601 STRATAGENE ™ pancreas (#937208) L0602 Pancreatic Islet pancreatic islet L0603 STRATAGENE ™ placenta (#937225) L0604 STRATAGENE ™ muscle muscle 937209 L0605 STRATAGENE ™ fetal fetal spleen spleen (#937205) L0606 NCI_CGAP_Lym5 follicular lymphoma L0607 NCI_CGAP_Lym6 mantle cell lymphoma L0608 STRATAGENE ™ lung lung carcinoma carcinoma 937218 L0609 Schiller astrocytoma astrocytoma L0611 Schiller meningioma meningioma L0612 Schiller oligodendroglioma oligodendroglioma L0615 22 week old human fetal liver cDNA library L0616 Chromosome 21 exon L0617 Chromosome 22 exon L0619 Chromosome 9 exon II L0622 HM1 L0623 HM3 pectoral muscle (after mastectomy) L0625 NCI_CGAP_AR1 bulk alveolar tumor L0626 NCI_CGAP_GC1 bulk germ cell seminoma L0627 NCI_CGAP_Co1 bulk tumor L0628 NCI_CGAP_Ov1 ovary bulk tumor L0629 NCI_CGAP_Mel3 metastatic melanoma to bowel L0630 NCI_CGAP_CNS1 substantia nigra L0631 NCI_CGAP_Br7 L0632 NCI_CGAP_Li5 hepatic adenoma L0634 NCI_CGAP_Ov8 serous adenocarcinoma L0635 NCI_CGAP_PNS1 dorsal root ganglion L0636 NCI_CGAP_Pit1 four pooled pituitary adenomas L0637 NCI_CGAP_Brn53 three pooled meningiomas L0638 NCI_CGAP_Brn35 tumor, 5 pooled (see description) L0639 NCI_CGAP_Brn52 tumor, 5 pooled (see description) L0640 NCI_CGAP_Br18 four pooled high-grade tumors, including two prima L0641 NCI_CGAP_Co17 juvenile granulosa tumor L0642 NCI_CGAP_Co18 moderately differentiated adenocarcinoma L0643 NCI_CGAP_Co19 moderately differentiated adenocarcinoma L0644 NCI_CGAP_Co20 moderately differentiated adenocarcinoma L0645 NCI_CGAP_Co21 moderately differentiated adenocarcinoma L0646 NCI_CGAP_Co14 moderately-differentiated adenocarcinoma L0647 NCI_CGAP_Sar4 five pooled sarcomas, including myxoid liposarcoma L0648 NCI_CGAP_Eso2 squamous cell carcinoma L0649 NCI_CGAP_GU1 2 pooled high-grade transitional cell tumors L0650 NCI_CGAP_Kid13 2 pooled Wilms'' tumors, one primary and one metast L0651 NCI_CGAP_Kid8 renal cell tumor L0652 NCI_CGAP_Lu27 four pooled poorly- differentiated adenocarcinomas L0653 NCI_CGAP_Lu28 two pooled squamous cell carcinomas L0654 NCI_CGAP_Lu31 L0655 NCI_CGAP_Lym12 lymphoma, follicular mixed small and large cell L0656 NCI_CGAP_Ov38 normal epithelium L0657 NCI_CGAP_Ov23 tumor, 5 pooled (see description) L0658 NCI_CGAP_Ov35 tumor, 5 pooled (see description) L0659 NCI_CGAP_Pan1 adenocarcinoma L0661 NCI_CGAP_Mel15 malignant melanoma, metastatic to lymph node L0662 NCI_CGAP_Gas4 poorly differentiated adenocarcinoma with signet r L0663 NCI_CGAP_Ut2 moderately-differentiated endometrial adenocarcino L0664 NCI_CGAP_Ut3 poorly-differentiated endometrial adenocarcinoma, L0665 NCI_CGAP_Ut4 serous papillary carcinoma, high grade, 2 pooled t L0666 NCI_CGAP_Ut1 well-differentiated endometrial adenocarcinoma, 7 L0667 NCI_CGAP_CML1 myeloid cells, 18 pooled CML cases, BCR/ABL rearra L0683 Stanley Frontal NS pool 2 frontal lobe (see description) L0686 Stanley Frontal SN pool 2 frontal lobe (see description) L0690 Testis, Subtracted L0697 Testis 1 L0698 Testis 2 L0708 NIH_MGC_17 rhabdomyosarcoma L0709 NIH_MGC_21 choriocarcinoma L0710 NIH_MGC_7 small cell carcinoma L0717 Gessler Wilms tumor L0731 Soares_pregnant_uterus_NbHPU L0738 Human colorectal cancer L0740 Soares melanocyte 2NbHM melanocyte L0741 Soares adult brain N2b4HB55Y L0742 Soares adult brain N2b5HB55Y L0743 Soares breast 2NbHBst L0744 Soares breast 3NbHBst L0745 Soares retina N2b4HR retina L0746 Soares retina N2b5HR retina L0747 Soares_fetal_heart_NbHH19W L0748 Soares fetal liver spleen 1NFLS L0749 Soares_fetal_liver_spleen_1NFLS_S1 L0750 Soares_fetal_lung_NbHL19W L0751 Soares ovary tumor NbHOT ovarian tumor L0752 Soares_parathyroid_tumor_NbHPA parathyroid tumor L0753 Soares_pineal_gland_N3HPG L0754 Soares placenta Nb2HP L0755 Soares_placenta_8to9weeks_2NbHP8to9W L0756 Soares_multiple_sclerosis_2NbHMSP multiple sclerosis lesions L0757 Soares_senescent_fibroblasts_NbHSF senescent fibroblast L0758 Soares_testis_NHT L0759 Soares_total_fetus_Nb2HF8_9w L0760 Barstead aorta HPLRB3 aorta L0761 NCI_CGAP_CLL1 B-cell, chronic lymphotic leukemia L0762 NCI_CGAP_Br1.1 breast L0763 NCI_CGAP_Br2 breast L0764 NCI_CGAP_Co3 colon L0765 NCI_CGAP_Co4 colon L0766 NCI_CGAP_GCB1 germinal center B cell L0767 NCI_CGAP_GC3 pooled germ cell tumors L0768 NCI_CGAP_GC4 pooled germ cell tumors L0769 NCI_CGAP_Brn25 anaplastic oligodendroglioma L0770 NCI_CGAP_Brn23 glioblastoma (pooled) L0771 NCI_CGAP_Co8 adenocarcinoma L0772 NCI_CGAP_Co10 colon tumor RER+ L0773 NCI_CGAP_Co9 colon tumor RER+ L0774 NCI_CGAP_Kid3 L0775 NCI_CGAP_Kid5 2 pooled tumors (clear cell type) L0776 NCI_CGAP_Lu5 carcinoid L0777 Soares_NhHMPu_S1 Pooled human melanocyte, fetal heart, and pregnant L0778 Barstead pancreas HPLRB1 L0779 Soares_NFL_T_GBC_S1 L0780 Soares_NSF_F8_9W_OT_PA_P_S1 L0782 NCI_CGAP_Pr21 normal prostate L0783 NCI_CGAP_Pr22 normal prostate L0784 NCI_CGAP_Lei2 leiomyosarcoma L0785 Barstead spleen HPLRB2 L0786 Soares_NbHFB L0787 NCI_CGAP_Sub1 L0788 NCI_CGAP_Sub2 L0789 NCI_CGAP_Sub3 L0790 NCI_CGAP_Sub4 L0791 NCI_CGAP_Sub5 L0792 NCI_CGAP_Sub6 L0793 NCI_CGAP_Sub7 L0794 NCI_CGAP_GC6 pooled germ cell tumors L0796 NCI_CGAP_Brn50 medulloblastoma L0800 NCI_CGAP_Co16 colon tumor, RER+ L0803 NCI_CGAP_Kid11 L0804 NCI_CGAP_Kid12 2 pooled tumors (clear cell type) L0805 NCI_CGAP_Lu24 carcinoid L0806 NCI_CGAP_Lu19 squamous cell carcinoma, poorly differentiated (4 L0807 NCI_CGAP_Ov18 fibrotheoma L0808 Barstead prostate BPH HPLRB4 1 L0809 NCI_CGAP_Pr28 L0879 BT0254 L0946 BT0333 L1057 BT0559 L1441 CT0249 L1446 CT0254 L1499 CT0322 L1651 HT0059 L1788 HT0229 L1819 HT0268 L1877 HT0340 L1878 HT0342 L2174 ST0240 L2251 Human fetal lung Fetal lung L2252 Human placenta placenta L2255 GLC corresponding non cancerous liver tissue L2257 NIH_MGC_65 adenocarcinoma L2258 NIH_MGC_67 retinoblastoma L2259 NIH_MGC_68 large cell carcinoma L2260 NIH_MGC_69 large cell carcinoma, undifferentiated L2261 NIH_MGC_70 epithelioid carcinoma L2262 NIH_MGC_72 melanotic melanoma L2263 NIH_MGC_66 adenocarcinoma L2264 NIH_MGC_71 leiomyosarcoma L2265 NIH_MGC_39 adenocarcinoma L2270 Lupski_dorsal_root_ganglion dorsal root ganglia L2281 BT0701 L2289 BT0757 L2333 CT0417 L2338 CT0432 L2346 CT0483 L2357 UT0021 L2367 UT0039 L2377 NN0054 L2380 NN0068 L2402 NN0118 L2412 NN0136 L2413 NN0141 L2467 NN1112 L2497 HT0618 L2498 HT0619 L2504 HT0636 L2518 HT0697 L2519 HT0698 L2522 HT0704 L2539 HT0727 L2543 HT0734 L2550 HT0743 L2570 HT0771 L2598 HT0809 L2599 HT0810 L2637 HT0877 L2640 HT0881 L2647 HT0894 L2650 HT0934 L2651 NIH_MGC_20 melanotic melanoma L2653 NIH_MGC_58 hypernephroma L2654 NIH_MGC_9 adenocarcinoma cell line L2655 NIH_MGC_55 from acute myelogenous leukemia L2657 NIH_MGC_54 from chronic myelogenous leukemia L2669 NT0022 L2670 NT0023 L2671 NT0024 L2677 NT0039 L2681 NT0048 L2686 NT0058 L2702 NT0098 L2708 NT0104 L2709 NT0105 L2738 GN0049 L2767 FT0044 L2791 FT0077 L2799 FT0096 L2800 FT0097 L2814 FT0128 L2817 FT0131 L2831 FT0162 L2842 UM0009 L2877 AN0027 L2884 AN0041 L2902 BN0036 L2904 BN0042 L2905 BN0046 L2906 BN0047 L2910 BN0070 L2915 BN0098 L2918 BN0114 L2919 BN0115 L2962 BN0221 L2991 BN0264 L2999 BN0273 L3002 BN0276 L3012 BN0296 L3058 EN0004 L3071 EN0026 L3081 ET0005 L3089 ET0018 L3104 ET0041 L3111 ET0058 L3117 ET0068 L3118 ET0070 L3119 ET0072 L3127 ET0084 L3144 MT0035 L3153 MT0049 L3154 MT0050 L3158 MT0057 L3199 OT0019 L3204 OT0034 L3207 OT0063 L3210 OT0067 L3216 OT0086 L3262 FN0073 L3278 FN0104 L3281 FN0107 L3311 FN0180 L3312 FN0181 L3316 FN0188 L3352 TN0027 L3357 TN0034 L3372 TN0068 L3374 TN0070 L3377 TN0079 L3378 TN0080 L3387 GKB hepatocellular carcinoma L3388 GKC hepatocellular carcinoma L3391 NIH_MGC_53 carcinoma, cell line L3403 AN0087 L3404 AN0089 L3421 BT0634 L3432 CT0461 L3435 CT0465 L3450 CT0508 L3459 FT0175 L3480 GN0057 L3484 GN0067 L3485 GN0070 L3491 GN0076 L3496 HT0572 L3499 HT0617 L3503 HT0870 L3504 HT0873 L3506 HT0879 L3511 HT0900 L3516 HT0913 L3518 HT0915 L3521 HT0919 L3530 HT0939 L3561 TN0025 L3562 TN0030 L3586 TN0120 L3603 UM0093 L3618 UT0050 L3631 UT0072 L3632 UT0074 L3642 ADA Adrenal gland L3643 ADB Adrenal gland L3644 ADC Adrenal gland L3645 Cu adrenal cortico adenoma for Cushing''s syndrome L3646 DCA L3649 DCB L3653 HTB Hypothalamus L3655 HTC Hypothalamus L3657 HTF Hypothalamus L3658 cdA pheochromocytoma L3659 CB cord blood L3661 NPA pituitary L3665 NIH_MGC_75 L3667 NIH_MGC_79 L3684 BT0812 L3705 CT0486 L3713 CT0524 L3722 GN0030 L3729 GN0079 L3744 HT0916 L3750 HT0945 L3783 TN0136 L3807 UT0077 L3808 UT0078 L3811 NPC pituitary L3812 NPD pituitary L3813 TP pituitary tumor L3814 BM Bone marrow L3815 MDS Bone marrow L3816 HEMBA1 whole embryo, mainly head L3817 HEMBB1 whole embryo, mainly body L3819 NIH_MGC_76 L3824 NT2RM2 L3825 NT2RM4 L3826 NT2RP1 L3827 NT2RP2 L3828 NT2RP3 L3829 NT2RP4 L3831 OVARC1 ovary, tumor tissue L3832 PLACE1 placenta L3833 PLACE2 placenta L3834 PLACE3 placenta L3837 THYRO1 thyroid gland L3839 Y79AA1 L3841 NIH_MGC_18 large cell carcinoma L3871 NIH_MGC_19 neuroblastoma L3872 NCI_CGAP_Skn1 L3904 NCI_CGAP_Brn64 glioblastoma with EGFR amplification L3905 NCI_CGAP_Brn67 anaplastic oligodendroglioma with 1p/19q loss L4501 NCI_CGAP_Sub8 L4537 NCI_CGAP_Thy7 follicular adenoma (benign lesion) L4556 NCI_CGAP_HN13 squamous cell carcinoma L4560 NCI_CGAP_Ut7 tumor L4669 NCI_CGAP_Ov41 serous papillary tumor L4747 NCI_CGAP_Brn41 oligodendroglioma L5286 NCI_CGAP_Thy10 medullary carcinoma L5564 NCI_CGAP_HN20 L5565 NCI_CGAP_Brn66 glioblastoma with probably TP53 mutation and witho L5566 NCI_CGAP_Brn70 anaplastic oligodendroglioma L5568 NCI_CGAP_HN21 nasopharyngeal carcinoma L5569 NCI_CGAP_HN17 normal epithelium L5574 NCI_CGAP_HN19 normal epithelium L5575 NCI_CGAP_Brn65 glioblastoma without EGFR amplification L5622 NCI_CGAP_Skn3 L5623 NCI_CGAP_Skn4 squamous cell carcinoma Code Organ Cell Line Disease Vector AR022 AR023 AR024 AR025 AR026 AR027 AR028 AR029 AR030 AR031 AR032 AR033 AR034 AR035 AR036 AR037 AR038 AR039 AR040 AR041 AR042 AR043 AR044 AR050 AR051 AR052 AR053 AR054 AR055 AR056 AR057 AR058 AR059 AR060 AR061 AR062 AR063 AR064 AR065 AR066 AR067 AR068 AR069 AR070 AR071 AR072 AR073 AR074 AR075 AR076 AR077 AR078 AR079 AR080 AR081 AR082 AR083 AR084 AR085 AR086 AR087 AR088 AR089 AR090 AR091 AR092 AR093 AR094 AR095 AR096 AR097 AR098 AR099 AR100 AR101 AR102 AR103 AR104 AR105 AR106 AR119 AR120 AR121 AR124 AR125 AR126 AR127 AR130 AR131 AR135 AR136 AR139 AR140 AR141 AR152 AR153 AR154 AR155 AR161 AR162 AR163 AR164 AR165 AR166 AR167 AR168 AR169 AR170 AR171 AR172 AR173 AR174 AR175 AR176 AR177 AR178 AR179 AR180 AR181 AR182 AR183 AR184 AR185 AR186 AR187 AR188 AR189 AR190 AR191 AR192 AR193 AR194 AR195 AR196 AR197 AR198 AR199 AR200 AR201 AR202 AR203 AR204 AR205 AR206 AR207 AR208 AR209 AR210 AR211 AR212 AR213 AR214 AR215 AR216 AR217 AR218 AR219 AR221 AR222 AR223 AR224 AR225 AR226 AR227 AR228 AR229 AR230 AR231 AR232 AR233 AR234 AR235 AR236 AR237 AR238 AR239 AR240 AR241 AR242 AR243 AR244 AR245 AR246 AR247 AR248 AR249 AR250 AR251 AR252 AR253 AR254 AR255 AR256 AR257 AR258 AR259 AR260 AR261 AR262 AR263 AR264 AR265 AR266 AR267 AR268 AR269 AR270 AR271 AR272 AR273 AR274 AR275 AR276 AR277 AR278 AR279 AR280 AR281 AR282 AR283 AR284 AR285 AR286 AR287 AR288 AR289 AR290 AR291 AR292 AR293 AR294 AR295 AR296 AR297 AR298 AR299 AR300 AR301 AR302 AR303 AR304 AR306 AR307 AR308 AR309 AR310 AR311 AR312 AR313 AR314 AR315 AR317 AR318 AR326 AR327 AR328 AR355 AR356 AR357 AR358 AR359 AR360 AR361 AR362 AR363 AR364 AR365 AR366 AR367 AR368 AR369 AR370 AR371 AR372 AR373 AR374 AR375 AR376 AR377 AR378 AR381 AR382 AR383 AR384 AR385 AR386 AR387 AR388 AR389 AR390 AR391 AR392 AR393 AR394 AR398 AR399 AR400 AR401 AR402 AR404 AR405 AR406 AR407 AR408 AR409 AR410 AR411 AR412 AR413 AR414 AR415 AR416 AR417 AR418 AR419 AR420 AR421 AR422 AR423 AR424 AR425 AR427 AR428 AR429 AR430 AR431 AR432 AR433 AR434 AR435 AR436 AR438 AR439 AR440 AR441 AR442 AR443 AR444 H0002 Heart Uni-ZAP XR H0004 Spleen Uni-ZAP XR H0008 Uni-ZAP XR H0009 Uni-ZAP XR H0011 Kidney Uni-ZAP XR H0012 Kidney Uni-ZAP XR H0013 Embryo Uni-ZAP XR H0014 Gall Bladder Uni-ZAP XR H0015 Gall Bladder Uni-ZAP XR H0016 peritoneum Uni-ZAP XR H0020 Brain Uni-ZAP XR H0022 Lambda ZAP II H0023 Uni-ZAP XR H0024 Lung Uni-ZAP XR H0026 Lambda ZAP II H0028 Ovary pBLUESCRIPT ™ H0030 Uni-ZAP XR H0031 Placenta Uni-ZAP XR H0032 Prostate Uni-ZAP XR H0033 Uni-ZAP XR H0036 Small Int. Uni-ZAP XR H0038 Testis Uni-ZAP XR H0039 Pancreas disease Uni-ZAP XR H0040 Testis disease Uni-ZAP XR H0041 Bone Uni-ZAP XR H0042 Lung Uni-ZAP XR H0044 eye Uni-ZAP XR H0045 Esophagus disease Uni-ZAP XR H0046 Uterus disease Uni-ZAP XR H0047 Liver Uni-ZAP XR H0048 Uni-ZAP XR H0050 Heart Uni-ZAP XR H0051 Brain Uni-ZAP XR H0052 Brain Uni-ZAP XR H0056 Umbilical Uni-ZAP XR vein H0057 Uni-ZAP XR H0058 Thymus disease Lambda ZAP II H0059 Uterus disease Lambda ZAP II H0060 Blood Cell Line pBLUESCRIPT ™ H0061 Blood Cell Line pBLUESCRIPT ™ H0063 Thymus Uni-ZAP XR H0065 Esophagus Uni-ZAP XR H0068 Skin disease Uni-ZAP XR H0069 Blood Cell Line Uni-ZAP XR H0070 Pancreas Uni-ZAP XR H0071 Adrenal Uni-ZAP XR gland H0073 Muscle disease Uni-ZAP XR H0075 Blood Cell Line Uni-ZAP XR H0076 Blood Cell Line Uni-ZAP XR H0078 Lung disease Lambda ZAP II H0081 Skin Uni-ZAP XR H0083 Uni-ZAP XR H0085 Lambda ZAP II H0086 Sk Muscle disease Uni-ZAP XR H0087 pBLUESCRIPT ™ H0090 T-Cell disease Uni-ZAP XR H0097 Heart pBLUESCRIPT ™ H0098 Liver Uni-ZAP XR H0099 Lung pBLUESCRIPT ™ H0100 Embryo Uni-ZAP XR H0101 Embryo Lambda ZAP II H0102 Embryo pBLUESCRIPT ™ H0103 Brain Uni-ZAP XR H0105 Heart pBLUESCRIPT ™ H0107 Adrenal pBLUESCRIPT ™ gland H0108 Lymph Uni-ZAP XR Node H0109 Blood Cell Line pBLUESCRIPT ™ H0110 Ovary pBLUESCRIPT ™ H0111 Placenta pBLUESCRIPT ™ H0116 Thymus pBLUESCRIPT ™ H0118 Kidney Uni-ZAP XR H0119 Kidney Uni-ZAP XR H0120 Spleen Uni-ZAP XR H0121 eye Uni-ZAP XR H0122 Sk Muscle Uni-ZAP XR H0123 Brain Uni-ZAP XR H0124 Sk Muscle disease Uni-ZAP XR H0125 Blood Cell Line Uni-ZAP XR H0128 Uni-ZAP XR H0130 Prostate Cell Line Uni-ZAP XR H0131 Prostate Cell Line Uni-ZAP XR H0132 Prostate Cell Line Uni-ZAP XR H0134 Blood Cell Line Uni-ZAP XR H0135 Synovium Uni-ZAP XR H0136 Blood Cell Line Uni-ZAP XR H0139 Blood Cell Line Uni-ZAP XR H0140 Blood Cell Line Uni-ZAP XR H0141 Blood Cell Line Uni-ZAP XR H0144 Embryo Uni-ZAP XR H0147 Liver Uni-ZAP XR H0149 Embryo Uni-ZAP XR H0150 Testis Uni-ZAP XR H0151 Liver Uni-ZAP XR H0154 Skin disease Uni-ZAP XR H0156 Adrenal disease Uni-ZAP XR Gland H0158 Blood Cell Line Uni-ZAP XR H0159 Blood Cell Line Uni-ZAP XR H0161 Blood Cell Line Uni-ZAP XR H0163 Synovium Uni-ZAP XR H0165 Prostate disease Uni-ZAP XR H0166 Prostate disease Uni-ZAP XR H0167 Blood Cell Line Uni-ZAP XR H0169 Prostate disease Uni-ZAP XR H0170 Embryo Uni-ZAP XR H0171 Embryo Uni-ZAP XR H0172 Brain Lambda ZAP II H0176 Breast Cell Line Uni-ZAP XR H0177 Breast Cell Line Uni-ZAP XR H0178 Brain Uni-ZAP XR H0179 Blood Cell Line Uni-ZAP XR H0180 Breast disease Uni-ZAP XR H0181 Breast disease Uni-ZAP XR H0182 Breast disease Uni-ZAP XR H0183 Colon disease Uni-ZAP XR H0184 Liver disease Lambda ZAP II H0187 Blood Cell Line Lambda ZAP II H0188 Breast Uni-ZAP XR H0189 Blood Cell Line Uni-ZAP XR H0190 Blood Cell Line Uni-ZAP XR H0192 Blood Cell Line Uni-ZAP XR H0194 Brain pBLUESCRIPT ™ H0196 Heart Uni-ZAP XR H0197 Liver Uni-ZAP XR H0199 Liver Uni-ZAP XR H0200 peritoneum Uni-ZAP XR H0201 Brain pBLUESCRIPT ™ H0202 Blood Cell Line Uni-ZAP XR H0204 Colon pBLUESCRIPT ™ H0205 Colon pBLUESCRIPT ™ H0207 Prostate Cell Line pBLUESCRIPT ™ H0208 Lung pBLUESCRIPT ™ H0209 Brain Uni-ZAP XR H0211 Prostate pBLUESCRIPT ™ H0212 Prostate pBLUESCRIPT ™ H0213 Uni-ZAP XR H0214 Blood Cell Line pBLUESCRIPT ™ H0215 Blood Cell Line pBLUESCRIPT ™ H0216 Blood Cell Line pBLUESCRIPT ™ H0217 Blood Cell Line pBLUESCRIPT ™ H0218 Blood Cell Line Uni-ZAP XR H0219 Blood Cell Line Uni-ZAP XR H0220 Blood Cell Line Uni-ZAP XR H0222 Blood Cell Line Uni-ZAP XR H0223 Blood Cell Line Uni-ZAP XR H0224 Blood Cell Line Uni-ZAP XR H0225 Blood Cell Line Uni-ZAP XR H0229 Brain Lambda ZAP II H0230 Heart disease Uni-ZAP XR H0231 pBLUESCRIPT ™ H0232 pBLUESCRIPT ™ H0234 Liver pBLUESCRIPT ™ H0235 Liver pBLUESCRIPT ™ H0239 Kidney disease Uni-ZAP XR H0240 Breast Cell Line Uni-ZAP XR H0241 Breast Cell Line Uni-ZAP XR H0242 Heart pBLUESCRIPT ™ H0244 Embryo Uni-ZAP XR H0247 Blood Cell Line Uni-ZAP XR H0249 Embryo Uni-ZAP XR H0250 Uni-ZAP XR H0251 Cartilage disease Uni-ZAP XR H0252 Bone disease Uni-ZAP XR H0253 Testis Uni-ZAP XR H0254 Lymph Uni-ZAP XR Node H0255 Lymph Lambda ZAP II Node H0256 Blood Cell Line Uni-ZAP XR H0257 Blood Cell Line Uni-ZAP XR H0261 Brain Uni-ZAP XR H0263 Colon disease Lambda ZAP II H0264 Tonsil Uni-ZAP XR H0265 Blood Cell Line Uni-ZAP XR H0266 Vein Cell Line Lambda ZAP II H0267 Vein Cell Line Lambda ZAP II H0268 Umbilical Cell Line Lambda ZAP II vein H0269 Umbilical Cell Line Lambda ZAP II vein H0270 Pancreas Uni-ZAP XR H0271 Blood Cell Line Uni-ZAP XR H0272 Tonsil Uni-ZAP XR H0274 Spleen Uni-ZAP XR H0275 Adrenal pBLUESCRIPT ™ gland H0280 cell line Cell Line ZAP Express H0281 Lymph Cell Line ZAP Express Node H0282 Breast Uni-ZAP XR H0284 Bone Cell Line Uni-ZAP XR H0286 Bone Cell Line Uni-ZAP XR H0288 Bone Cell Line Uni-ZAP XR H0290 Bone Cell Line Uni-ZAP XR H0292 Bone Cell Line Uni-ZAP XR H0293 Uni-ZAP XR H0294 Placenta Cell Line Uni-ZAP XR H0295 Placenta Cell Line Uni-ZAP XR H0298 Breast Cell Line Uni-ZAP XR H0305 Cord Blood ZAP Express H0306 Cord Blood ZAP Express H0309 Synovium disease Uni-ZAP XR H0310 Brain Uni-ZAP XR H0316 Stomach Uni-ZAP XR H0318 Lymph disease Uni-ZAP XR Node H0320 Brain Uni-ZAP XR H0321 Nerve disease Uni-ZAP XR H0327 Brain Uni-ZAP XR H0328 Ovary disease Uni-ZAP XR H0329 Skin disease Uni-ZAP XR H0331 Liver disease Lambda ZAP II H0333 Blood vessel disease Lambda ZAP II H0334 Kidney disease Uni-ZAP XR H0339 Uni-ZAP XR H0341 Bone Cell Line Uni-ZAP XR Marrow H0343 disease Uni-ZAP XR H0344 Uni-ZAP XR H0345 Skin Uni-ZAP XR H0346 Brain disease Uni-ZAP XR H0349 Liver pCMVSport 1 H0350 Liver Uni-ZAP XR H0351 Brain disease Uni-ZAP XR H0352 disease Uni-ZAP XR H0354 Blood Cell Line pCMVSport 1 H0355 pCMVSport 1 H0356 Kidney pCMVSport 1 H0357 Liver Uni-ZAP XR H0359 ZAP Express H0360 disease H0361 disease pBLUESCRIPT ™ H0362 pSport1 H0366 ZAP Express H0369 Uni-ZAP XR H0370 disease Uni-ZAP XR H0371 disease Uni-ZAP XR H0372 Testis pCMVSport 1 H0373 Heart pCMVSport 1 H0374 pCMVSport 1 H0375 pCMVSport 1 H0376 Spleen pCMVSport 1 H0379 pSport1 H0380 pSport1 H0381 disease Uni-ZAP XR H0383 Uni-ZAP XR H0384 pCMVSport 1 H0386 Blood Cell Line pCMVSport 1 H0388 disease pBLUESCRIPT ™ H0390 disease pBLUESCRIPT ™ H0391 brain pSport1 H0392 brain pSport1 H0393 Liver pBLUESCRIPT ™ H0394 ZAP Express H0395 ZAP Express H0396 ZAP Express H0399 Lambda ZAP II H0400 Brain Lambda ZAP II H0402 Cord Blood ZAP Express H0403 Umbilical Cell Line Uni-ZAP XR vein H0404 Umbilical Cell Line Uni-ZAP XR vein H0405 pBLUESCRIPT ™ H0406 Uni-ZAP XR H0408 pBLUESCRIPT ™ H0409 Brain pBLUESCRIPT ™ H0410 Bladder pSport1 H0411 Bladder pSport1 H0412 Umbilical Cell Line pSport1 vein H0413 Umbilical Cell Line pSport1 vein H0414 Ovary disease pSport1 H0415 Ovary disease pCMVSport 2.0 H0416 Blood Cell Line pBLUESCRIPT ™ H0417 pBLUESCRIPT ™ H0418 pBLUESCRIPT ™ H0419 Uni-ZAP XR H0421 pBLUESCRIPT ™ H0422 Blood Cell Line pSport1 H0423 Blood Cell Line pSport1 H0424 pBLUESCRIPT ™ H0427 pSport1 H0428 Ovary pSport1 H0429 cell line Cell Line ZAP Express H0431 Kidney pBLUESCRIPT ™ H0433 Umbilical Cell Line pBLUESCRIPT ™ vein H0434 pBLUESCRIPT ™ H0435 Ovary pCMVSport 2.0 H0436 Blood Cell Line pSport1 H0437 Umbilical Cell Line Lambda ZAP II vein H0438 ZAP Express H0439 pBLUESCRIPT ™ H0441 Kidney pBLUESCRIPT ™ H0443 pSport1 H0444 Spleen disease pSport1 H0445 Spleen disease pSport1 H0449 pSport1 H0453 Kidney pBLUESCRIPT ™ H0455 Brain pBLUESCRIPT ™ H0457 pSport1 H0458 pSport1 H0459 pCMVSport 2.0 H0462 Brain pBLUESCRIPT ™ H0477 Tonsil pSport1 H0478 Salivary pSport1 gland H0479 Salivary pSport1 gland H0483 pSport1 H0484 pSport1 H0485 disease pCMVSport 2.0 H0486 disease pCMVSport 2.0 H0487 pCMVSport 2.0 H0488 pCMVSport 2.0 H0489 Intestine disease pSport1 H0490 Blood Cell Line Uni-ZAP XR H0491 Blood Cell Line Uni-ZAP XR H0492 Blood Cell Line Uni-ZAP XR H0494 pCMVSport 2.0 H0497 HEL 92.1.7 pSport1 H0505 pSport1 H0506 Colon pSport1 H0509 Liver disease pCMVSport 3.0 H0510 Liver pCMVSport 3.0 H0512 pCMVSport 2.0 H0518 pCMVSport 3.0 H0519 pCMVSport 3.0 H0520 pSport1 H0521 pCMVSport 3.0 H0522 pCMVSport 3.0 H0525 PCRII H0528 pCMVSport 3.0 H0529 pCMVSport 3.0 H0530 pSport1 H0538 Lymph node pSport1 H0539 Pancreas disease pSport1 H0540 Skin pSport1 H0542 pCMVSport 3.0 H0543 pCMVSport 3.0 H0544 pCMVSport 3.0 H0545 pCMVSport 3.0 H0546 pCMVSport 3.0 H0547 pSport1 H0549 Uni-ZAP XR H0550 Uni-ZAP XR H0551 pCMVSport 3.0 H0553 pCMVSport 3.0 H0555 Kidney disease pCMVSport 3.0 H0556 Blood Cell Line Uni-ZAP XR H0559 Blood Cell Line Uni-ZAP XR H0560 pCMVSport 3.0 H0561 pCMVSport 3.0 H0562 pCMVSport 2.0 H0563 pCMVSport 2.0 H0564 pCMVSport 2.0 H0566 pCMVSport 2.0 H0567 pCMVSport 2.0 H0569 pCMVSport 2.0 H0570 pCMVSport 2.0 H0571 pCMVSport 2.0 H0572 pCMVSport 2.0 H0574 Liver disease Lambda ZAP II H0575 Lung Uni-ZAP XR H0576 Blood Cell Line Lambda ZAP II H0579 Heart pSport1 H0580 pCMVSport 3.0 H0581 Bone pCMVSport 3.0 Marrow H0583 B Cell disease pCMVSport 3.0 H0584 Blood Cell Line Uni-ZAP XR H0585 Blood Cell Line Uni-ZAP XR H0586 groin disease pCMVSport 3.0 H0587 groin disease pCMVSport 3.0 H0589 Cord Blood ZAP Express H0590 Small Int. Uni-ZAP XR H0591 T-Cell disease Uni-ZAP XR H0592 disease pCMVSport 3.0 H0593 pCMVSport 3.0 H0594 Lung disease Lambda ZAP II H0595 disease Uni-ZAP XR H0596 Colon Lambda ZAP II H0597 Lambda ZAP II H0598 Stomach Uni-ZAP XR H0599 Heart Uni-ZAP XR H0600 disease pCMVSport 3.0 H0601 disease pCMVSport 3.0 H0602 disease pCMVSport 3.0 H0604 pBLUESCRIPT ™ H0606 Breast disease Uni-ZAP XR H0607 pCMVSport 1 H0609 pCMVSport 1 H0610 pCMVSport 1 H0611 pCMVSport 1 H0613 pCMVSport 1 H0614 pCMVSport 1 H0615 Ovary disease Uni-ZAP XR H0616 Testis Uni-ZAP XR H0617 Breast disease Uni-ZAP XR H0618 Testis Uni-ZAP XR H0619 Heart Uni-ZAP XR H0620 Kidney Uni-ZAP XR H0622 Pancreas disease Uni-ZAP XR H0623 Umbilical Uni-ZAP XR vein H0624 Embryo Uni-ZAP XR H0625 pSport1 H0626 pSport1 H0627 pSport1 H0628 Uni-ZAP XR H0629 pCMVSport 1 H0631 pSport1 H0632 Liver Lambda ZAP II H0633 disease pSport1 H0634 Testis disease Uni-ZAP XR H0635 Blood Cell Line Uni-ZAP XR H0637 pSport1 H0638 pSport1 H0640 Other H0641 pSport1 H0642 Other H0643 Other H0644 Placenta Uni-ZAP XR H0645 Heart Uni-ZAP XR H0646 pSport1 H0647 disease pSport1 H0648 disease pSport1 H0649 pSport1 H0650 pCMVSport 3.0 H0651 pSport1 H0652 pSport1 H0653 pSport1 H0654 Other H0656 pSport1 H0657 pSport1 H0658 Ovary & disease pSport1 Fallopian Tubes H0659 Ovary disease pSport1 H0660 disease pSport1 H0661 disease pSport1 H0662 Breast pSport1 H0663 Breast disease pSport1 H0664 Breast disease pSport1 H0665 pSport1 H0666 disease pSport1 H0667 pSport1 H0668 pSport1 H0670 pSport1 H0671 pSport1 H0672 Ovary pSport1 H0673 Prostate Uni-ZAP XR H0674 Prostate Uni-ZAP XR H0675 pCMVSport 3.0 H0676 pCMVSport 3.0 H0677 PCRII H0678 Placenta Other H0682 pCMVSport 3.0 H0683 pCMVSport 3.0 H0684 Ovaries pCMVSport 3.0 H0685 pCMVSport 3.0 H0686 pCMVSport 3.0 H0687 Ovary pCMVSport 3.0 H0688 pCMVSport 3.0 H0689 pCMVSport 3.0 H0690 pCMVSport 3.0 H0691 pCMVSport 3.0 H0693 pCMVSport 3.0 H0694 prostate pCMVSport 3.0 gland H0695 pCMVSport 3.0 N0003 N0006 N0007 N0009 S0001 Brain Lambda ZAP II S0002 blood Cell Line Uni-ZAP XR S0003 bone disease Uni-ZAP XR S0004 Prostate Lambda ZAP II S0006 disease pCDNA S0007 Uni-ZAP XR S0010 Uni-ZAP XR S0011 bone disease Uni-ZAP XR S0013 prostate Uni-ZAP XR S0014 Kidney Uni-ZAP XR S0015 Kidney Uni-ZAP XR S0016 Kidney Uni-ZAP XR S0021 Brain ZAP Express S0022 Uni-ZAP XR S0023 S0024 S0026 Bone Cell Line Uni-ZAP XR marrow S0027 Pulmanary Cell Line Uni-ZAP XR artery S0028 Pulmanary Cell Line Uni-ZAP XR artery S0029 brain Uni-ZAP XR S0030 Brain Uni-ZAP XR S0031 spinal cord Uni-ZAP XR S0032 Pulmanary Cell Line Uni-ZAP XR artery S0035 Brain Uni-ZAP XR S0036 Uni-ZAP XR S0037 Pulmanary Cell Line Uni-ZAP XR artery S0038 ZAP Express S0039 Brain Uni-ZAP XR S0040 Uni-ZAP XR S0042 ZAP Express S0044 Prostate disease Uni-ZAP XR S0045 endothelial Cell Line Uni-ZAP XR cell-lung S0046 endothelial Cell Line Uni-ZAP XR cell-lung S0048 disease Uni-ZAP XR S0049 Uni-ZAP XR S0050 disease Uni-ZAP XR S0051 disease Uni-ZAP XR S0052 blood Cell Line Uni-ZAP XR S0053 blood Cell Line Uni-ZAP XR S0106 BRAIN disease Uni-ZAP XR S0110 Brain disease Uni-ZAP XR S0112 Brain Uni-ZAP XR S0114 Cell Line Uni-ZAP XR S0116 Bone Uni-ZAP XR marrow S0122 bone disease pBLUESCRIPT ™ S0124 Pulmanary Cell Line Uni-ZAP XR artery S0126 Knee Cell Line Uni-ZAP XR S0132 Uni-ZAP XR S0134 Cell Line Uni-ZAP XR S0136 Bone Cell Line Lambda ZAP II marrow S0140 lung Cell Line Uni-ZAP XR S0142 blood Cell Line Uni-ZAP XR S0144 Uni-ZAP XR S0146 Prostate Uni-ZAP XR S0148 prostate Uni-ZAP XR S0150 Prostate Cell Line Uni-ZAP XR S0152 Uni-ZAP XR S0168 pBLUESCRIPT ™ S0176 prostate Uni-ZAP XR S0180 disease Uni-ZAP XR S0182 Uni-ZAP XR S0188 disease pSport1 S0190 pSport1 S0192 pSport1 S0194 pSport1 S0196 pSport1 S0206 Pulmanary Cell Line pBLUESCRIPT ™ artery S0208 pSport1 S0210 pSport1 S0212 pSport1 S0214 bone disease Uni-ZAP XR S0216 blood Cell Line Uni-ZAP XR S0218 Cell Line Uni-ZAP XR S0220 Brain ZAP Express S0222 Brain disease Uni-ZAP XR S0242 pSport1 S0250 Femur disease pCMVSport 2.0 S0260 spinal cord Uni-ZAP XR S0276 Synovial pSport1 tissue S0278 Uni-ZAP XR S0280 Uni-ZAP XR S0282 Brain Lambda ZAP II S0292 Bone disease pSport1 S0294 Larynx, vocal disease pSport1 cord S0298 Bone pSport1 marrow S0300 Brain Uni-ZAP XR S0306 pSport1 S0308 pSport1 S0310 pSport1 S0312 disease pSport1 S0314 disease pSport1 S0316 pSport1 S0318 pSport1 S0328 Uvula disease pSport1 S0330 Uvula pSport1 S0332 Hypopharynx pSport1 S0334 pSport1 S0336 pSport1 S0338 disease pSport1 S0340 disease pSport1 S0342 Uni-ZAP XR S0344 blood Cell Line Uni-ZAP XR S0346 Uni-ZAP XR S0348 disease pSport1 S0350 Hypopharynx disease pSport1 S0352 disease pSport1 S0354 Colon pSport1 S0356 Colon disease pSport1 S0358 Colon pSport1 S0360 Colon disease pSport1 S0362 pSport1 S0364 pSport1 S0366 pSport1 S0368 pSport1 S0370 disease pSport1 S0372 disease pSport1 S0374 pSport1 S0376 disease pSport1 S0378 pSport1 S0380 disease pSport1 S0382 disease pSport1 S0384 disease pSport1 S0386 Brain ZAP Express S0388 disease Uni-ZAP XR S0390 Pulmanary Cell Line Uni-ZAP XR artery S0392 pSport1 S0394 pSport1 S0398 pSport1 S0400 pSport1 S0402 pSport1 S0404 pSport1 S0406 pSport1 S0408 pSport1 S0410 pSport1 S0412 disease Other S0414 Other S0418 pCMVSport 3.0 S0420 pSport1 S0422 pCMVSport 3.0 S0424 pSport1 S0426 blood Cell Line Uni-ZAP XR S0428 blood Cell Line Uni-ZAP XR S0430 pSport1 S0432 pSport1 S0434 disease pSport1 S0436 disease pSport1 S0438 pSport1 S0440 pSport1 S0442 pSport1 S0444 disease pSport1 S0446 pSport1 S0448 pSport1 S0450 pSport1 S0452 pSport1 S0454 Placenta pSport1 S0456 pSport1 S0458 pSport1 S0460 pSport1 S0462 pSport1 S0464 pSport1 S0466 disease pSport1 S0468 pSport1 S0470 disease pSport1 S0472 pSport1 S0474 Blood Other platelets S0665 Uni-ZAP XR S3012 Pulmanary Cell Line pBLUESCRIPT ™ artery S3014 Pulmanary Cell Line pBLUESCRIPT ™ artery S6014 Brain ZAP Express S6016 Brain disease Uni-ZAP XR S6022 Uni-ZAP XR S6024 Brain disease Uni-ZAP XR S6026 Brain Uni-ZAP XR S6028 Brain disease Uni-ZAP XR T0002 Blood Cell Line pBLUESCRIPT ™ SK− T0003 pBLUESCRIPT ™ SK− T0004 pBLUESCRIPT ™ SK− T0006 pBLUESCRIPT ™ SK− T0008 disease pBLUESCRIPT ™ SK− T0010 Other T0023 disease pBLUESCRIPT ™ SK− T0039 pBLUESCRIPT ™ SK− T0040 pBLUESCRIPT ™ SK− T0041 pBLUESCRIPT ™ SK− T0042 pBLUESCRIPT ™ SK− T0048 pBLUESCRIPT ™ SK− T0049 pBLUESCRIPT ™ SK− T0060 pBLUESCRIPT ™ SK− T0067 pBLUESCRIPT ™ SK− T0068 pBLUESCRIPT ™ SK− T0069 pBLUESCRIPT ™ SK− T0071 pBLUESCRIPT ™ SK− T0079 pBLUESCRIPT ™ SK− T0082 pBLUESCRIPT ™ SK− T0086 disease pBLUESCRIPT ™ SK− T0103 pBLUESCRIPT ™ SK− T0104 pBLUESCRIPT ™ SK− T0109 pBLUESCRIPT ™ SK− T0110 pBLUESCRIPT ™ SK− T0112 pBLUESCRIPT ™ SK− T0114 pBLUESCRIPT ™ SK− T0115 pBLUESCRIPT ™ SK− L0002 L0005 L0015 L0021 L0022 L0024 L0040 L0041 L0045 L0053 L0055 L0065 L0070 L0096 L0097 L0103 L0105 L0142 L0143 L0151 L0157 brain L0163 heart L0182 HeLa L0187 HT1080 L0194 Patu 8988t L0295 liver L0309 E8CASS; variant of MCF7 L0351 BA, M13-derived L0352 BA, M13-derived L0355 Bluescript L0356 Bluescript L0361 ovary Bluescript SK L0362 Bluescript SK− L0363 Bluescript SK− L0364 Bluescript SK− L0365 Bluescript SK− L0366 Bluescript SK− L0367 Bluescript SK− L0368 Bluescript SK− L0369 adrenal Bluescript SK− gland L0370 brain Bluescript SK− L0371 breast Bluescript SK− L0372 colon Bluescript SK− L0373 colon Bluescript SK− L0374 colon Bluescript SK− L0375 kidney Bluescript SK− L0376 larynx Bluescript SK− L0378 lung Bluescript SK− L0379 lymph node Bluescript SK− L0381 pharynx Bluescript SK− L0382 prostate Bluescript SK− L0383 prostate Bluescript SK− L0384 prostate Bluescript SK− L0385 stomach Bluescript SK− L0386 tongue Bluescript SK− L0387 tonsil Bluescript SK− L0388 Bluescript SK− L0389 Bluescript SK− L0394 gt11 L0404 Lafmid A L0411 Lafmid BA L0415 Lafmid BA L0418 Lafmid BA L0428 Lafmid BA L0434 lafmid BA L0435 lafmid BA L0438 brain lafmid BA L0439 whole brain Lafmid BA L0443 Lafmid BK L0446 Lafmid BK L0454 lambda gt10 L0455 eye lambda gt10 L0456 eye lambda gt10 L0457 pooled lambda gt10 L0459 Lambda gt11 L0460 Lambda gt11 L0462 lambda gt11 L0463 brain lambda gt11 L0465 lambda nm1149 L0468 lambda zap L0471 Lambda ZAP Express L0475 KG1-a Lambda Zap Express (STRATAGENE ™) L0476 Lambda ZAP II L0480 Lambda ZAP, pBLUESCRIPT ™ SK(−) L0481 Lambda ZAPII L0483 Lambda ZAPII L0485 leg muscle Lambda ZAPII L0492 pAMP L0493 ovary pAMP1 L0497 bone pAMP1 marrow L0498 bone pAMP1 marrow L0499 bone pAMP1 marrow L0500 brain pAMP1 L0502 breast pAMP1 L0503 breast pAMP1 L0504 breast pAMP1 L0505 breast pAMP1 L0506 breast pAMP1 L0507 breast pAMP1 L0508 lung pAMP1 L0509 lung pAMP1 L0511 ovary pAMP1 L0512 ovary pAMP1 L0513 ovary pAMP1 L0514 ovary pAMP1 L0515 ovary pAMP1 L0517 pAMP10 L0518 pAMP10 L0519 pAMP10 L0520 pAMP10 L0521 pAMP10 L0522 pAMP10 L0523 pAMP10 L0524 pAMP10 L0525 pAMP10 L0526 pAMP10 L0527 pAMP10 L0528 pAMP10 L0529 pAMP10 L0530 pAMP10 L0532 pAMP10 L0533 bone pAMP10 marrow L0534 brain pAMP10 L0536 breast pAMP10 L0539 placenta pAMP10 L0540 prostate pAMP10 L0542 prostate pAMP10 L0543 prostate pAMP10 L0544 prostate pAMP10 L0545 prostate pAMP10 L0546 prostate pAMP10 L0547 prostate pAMP10 L0549 pAMP10 L0550 pAMP10 L0551 pAMP10 L0554 liver pAMP10 L0558 ovary pAMP10 L0559 ovary pAMP10 L0560 tongue pAMP10 L0561 tongue pAMP10 L0562 HeLa cell pAMP10 line; ATCC ™ L0564 pBLUESCRIPT ™ L0565 Hip pBLUESCRIPT ™ L0581 liver pBLUESCRIPT ™ SK L0584 pBLUESCRIPT ™ SK(+) L0586 pBLUESCRIPT ™ SK(−) L0587 pBLUESCRIPT ™ SK− L0588 pBLUESCRIPT ™ SK− L0589 pBLUESCRIPT ™ SK− L0590 pBLUESCRIPT ™ SK− L0591 pBLUESCRIPT ™ SK− L0592 pBLUESCRIPT ™ SK− L0593 pBLUESCRIPT ™ SK− L0594 pBLUESCRIPT ™ SK− L0595 brain pBLUESCRIPT ™ SK− L0596 colon pBLUESCRIPT ™ SK− L0597 cornea pBLUESCRIPT ™ SK− L0598 ear pBLUESCRIPT ™ SK− L0599 lung pBLUESCRIPT ™ SK− L0600 nose pBLUESCRIPT ™ SK− L0601 pancreas pBLUESCRIPT ™ SK− L0602 pancreas pBLUESCRIPT ™ SK− L0603 placenta pBLUESCRIPT ™ SK− L0604 skeletal pBLUESCRIPT ™ SK− muscle L0605 spleen pBLUESCRIPT ™ SK− L0606 lymph node pBLUESCRIPT ™ SK− L0607 lymph node pBLUESCRIPT ™ SK− L0608 lung NCI-H69 pBLUESCRIPT ™ SK− L0609 brain pBLUESCRIPT ™ SK− (STRATAGENE ™) L0611 brain pBLUESCRIPT ™ SK− (STRATAGENE ™) L0612 brain pBLUESCRIPT ™ SK− (STRATAGENE ™) L0615 pBLUESCRIPT ™ II SK(−) L0616 pBLUESCRIPT ™ II KS+ L0617 pBLUESCRIPT ™ II KS+ L0619 pBLUESCRIPT ™ II KS+ L0622 pcDNAII (Invitrogen) L0623 pcDNAII (Invitrogen) L0625 pCMV-SPORT2 L0626 pCMV-SPORT2 L0627 colon pCMV-SPORT2 L0628 ovary pCMV-SPORT2 L0629 bowel (skin pCMV-SPORT4 primary) L0630 brain pCMV-SPORT4 L0631 breast pCMV-SPORT4 L0632 liver pCMV-SPORT4 L0634 ovary pCMV-SPORT4 L0635 peripheral pCMV-SPORT4 nervous system L0636 brain pCMV-SPORT6 L0637 brain pCMV-SPORT6 L0638 brain pCMV-SPORT6 L0639 brain pCMV-SPORT6 L0640 breast pCMV-SPORT6 L0641 colon pCMV-SPORT6 L0642 colon pCMV-SPORT6 L0643 colon pCMV-SPORT6 L0644 colon pCMV-SPORT6 L0645 colon pCMV-SPORT6 L0646 colon pCMV-SPORT6 L0647 connective pCMV-SPORT6 tissue L0648 esophagus pCMV-SPORT6 L0649 genitourinary pCMV-SPORT6 tract L0650 kidney pCMV-SPORT6 L0651 kidney pCMV-SPORT6 L0652 lung pCMV-SPORT6 L0653 lung pCMV-SPORT6 L0654 lung, cell pCMV-SPORT6 line L0655 lymph node pCMV-SPORT6 L0656 ovary pCMV-SPORT6 L0657 ovary pCMV-SPORT6 L0658 ovary pCMV-SPORT6 L0659 pancreas pCMV-SPORT6 L0661 skin pCMV-SPORT6 L0662 stomach pCMV-SPORT6 L0663 uterus pCMV-SPORT6 L0664 uterus pCMV-SPORT6 L0665 uterus pCMV-SPORT6 L0666 uterus pCMV-SPORT6 L0667 whole blood pCMV-SPORT6 L0683 brain pCR2.1-TOPO (Invitrogen) L0686 brain pCR2.1-TOPO (Invitrogen) L0690 pCRII L0697 PGEM 5zf(+) L0698 PGEM 5zf(+) L0708 muscle pOTB7 L0709 placenta pOTB7 L0710 lung MGC3 pOTB7 L0717 pSPORT1 L0731 uterus pT7T3-Pac L0738 pT7T3D L0740 pT7T3D (PHARMACIA ™) with a modified polylinker L0741 brain pT7T3D (PHARMACIA ™) with a modified polylinker L0742 brain pT7T3D (PHARMACIA ™) with a modified polylinker L0743 breast pT7T3D (PHARMACIA ™) with a modified polylinker L0744 breast pT7T3D (PHARMACIA ™) with a modified polylinker L0745 eye pT7T3D (PHARMACIA ™) with a modified polylinker L0746 eye pT7T3D (PHARMACIA ™) with a modified polylinker L0747 heart pT7T3D (PHARMACIA ™) with a modified polylinker L0748 Liver and pT7T3D Spleen (PHARMACIA ™) with a modified polylinker L0749 Liver and pT7T3D Spleen (PHARMACIA ™) with a modified polylinker L0750 lung pT7T3D (PHARMACIA ™) with a modified polylinker L0751 ovary pT7T3D (PHARMACIA ™) with a modified polylinker L0752 parathyroid pT7T3D gland (PHARMACIA ™) with a modified polylinker L0753 pineal gland pT7T3D (PHARMACIA ™) with a modified polylinker L0754 placenta pT7T3D (PHARMACIA ™) with a modified polylinker L0755 placenta pT7T3D (PHARMACIA ™) with a modified polylinker L0756 pT7T3D (PHARMACIA ™) with a modified polylinker V_TYPE L0757 pT7T3D (PHARMACIA ™) with a modified polylinker V_TYPE L0758 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0759 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0760 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0761 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0762 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0763 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0764 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0765 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0766 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0767 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0768 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0769 brain pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0770 brain pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0771 colon pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0772 colon pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0773 colon pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0774 kidney pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0775 kidney pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0776 lung pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0777 mixed (see pT7T3D-Pac below) (PHARMACIA ™) with a modified polylinker L0778 pancreas pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0779 pooled pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0780 pooled pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0782 prostate pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0783 prostate pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0784 soft tissue pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0785 spleen pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0786 whole brain pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0787 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0788 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0789 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0790 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0791 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0792 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0793 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0794 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0796 brain pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0800 colon pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0803 kidney pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0804 kidney pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0805 lung pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0806 lung pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0807 ovary pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0808 prostate pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0809 prostate pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L0879 breast puc18 L0946 breast puc18 L1057 breast puc18 L1441 colon puc18 L1446 colon puc18 L1499 colon puc18 L1651 head_neck puc18 L1788 head_neck puc18 L1819 head_neck puc18 L1877 head_neck puc18 L1878 head_neck puc18 L2174 stomach puc18 L2251 L2252 L2255 pBLUESCRIPT ™ sk(−) L2257 colon pCMV-SPORT6 L2258 eye pCMV-SPORT6 L2259 lung pCMV-SPORT6 L2260 lung pCMV-SPORT6 L2261 pancreas pCMV-SPORT6 L2262 skin pCMV-SPORT6 L2263 ovary pCMV-SPORT6 L2264 uterus pCMV-SPORT6 L2265 pancreas pOTB7 L2270 pCMV-SPORT6 (Life Technologies) L2281 breast puc18 L2289 breast puc18 L2333 colon puc18 L2338 colon puc18 L2346 colon puc18 L2357 uterus_tumor puc18 L2367 uterus_tumor puc18 L2377 nervous_normal puc18 L2380 nervous_normal puc18 L2402 nervous_normal puc18 L2412 nervous_normal puc18 L2413 nervous_normal puc18 L2467 nervous_normal puc18 L2497 head_neck puc18 L2498 head_neck puc18 L2504 head_neck puc18 L2518 head_neck puc18 L2519 head_neck puc18 L2522 head_neck puc18 L2539 head_neck puc18 L2543 head_neck puc18 L2550 head_neck puc18 L2570 head_neck puc18 L2598 head_neck puc18 L2599 head_neck puc18 L2637 head_neck puc18 L2640 head_neck puc18 L2647 head_neck puc18 L2650 head_neck puc18 L2651 skin pOTB7 L2653 kidney pDNR-LIB (CLONTECH ™) L2654 ovary pOTB7 L2655 bone pDNR-LIB marrow (CLONTECH ™) L2657 bone pDNR-LIB marrow (CLONTECH ™) L2669 nervous_tumor puc18 L2670 nervous_tumor puc18 L2671 nervous_tumor puc18 L2677 nervous_tumor puc18 L2681 nervous_tumor puc18 L2686 nervous_tumor puc18 L2702 nervous_tumor puc18 L2708 nervous_tumor puc18 L2709 nervous_tumor puc18 L2738 placenta_normal puc18 L2767 prostate_tumor puc18 L2791 prostate_tumor puc18 L2799 prostate_tumor puc18 L2800 prostate_tumor puc18 L2814 prostate_tumor puc18 L2817 prostate_tumor puc18 L2831 prostate_tumor puc18 L2842 uterus puc18 L2877 amnion_normal puc18 L2884 amnion_normal puc18 L2902 breast_normal puc18 L2904 breast_normal puc18 L2905 breast_normal puc18 L2906 breast_normal puc18 L2910 breast_normal puc18 L2915 breast_normal puc18 L2918 breast_normal puc18 L2919 breast_normal puc18 L2962 breast_normal puc18 L2991 breast_normal puc18 L2999 breast_normal puc18 L3002 breast_normal puc18 L3012 breast_normal puc18 L3058 lung_normal puc18 L3071 lung_normal puc18 L3081 lung_tumor puc18 L3089 lung_tumor puc18 L3104 lung_tumor puc18 L3111 lung_tumor puc18 L3117 lung_tumor puc18 L3118 lung_tumor puc18 L3119 lung_tumor puc18 L3127 lung_tumor puc18 L3144 marrow puc18 L3153 marrow puc18 L3154 marrow puc18 L3158 marrow puc18 L3199 ovary puc18 L3204 ovary puc18 L3207 ovary puc18 L3210 ovary puc18 L3216 ovary puc18 L3262 prostate_normal puc18 L3278 prostate_normal puc18 L3281 prostate_normal puc18 L3311 prostate_normal puc18 L3312 prostate_normal puc18 L3316 prostate_normal puc18 L3352 testis_normal puc18 L3357 testis_normal puc18 L3372 testis_normal puc18 L3374 testis_normal puc18 L3377 testis_normal puc18 L3378 testis_normal puc18 L3387 pBLUESCRIPT ™ sk(−) L3388 pBLUESCRIPT ™ sk(−) L3391 bladder pDNR-LIB (CLONTECH ™) L3403 amnion_normal puc18 L3404 amnion_normal puc18 L3421 breast puc18 L3432 colon puc18 L3435 colon puc18 L3450 colon puc18 L3459 prostate_tumor puc18 L3480 placenta_normal puc18 L3484 placenta_normal puc18 L3485 placenta_normal puc18 L3491 placenta_normal puc18 L3496 head_neck puc18 L3499 head_neck puc18 L3503 head_neck puc18 L3504 head_neck puc18 L3506 head_neck puc18 L3511 head_neck puc18 L3516 head_neck puc18 L3518 head_neck puc18 L3521 head_neck puc18 L3530 head_neck puc18 L3561 testis_normal puc18 L3562 testis_normal puc18 L3586 testis_normal puc18 L3603 uterus puc18 L3618 uterus_tumor puc18 L3631 uterus_tumor puc18 L3632 uterus_tumor puc18 L3642 pBLUESCRIPT ™ sk(−) L3643 pBLUESCRIPT ™ sk(−) L3644 pBLUESCRIPT ™ sk(−) L3645 pBLUESCRIPT ™ sk(−) L3646 pTriplEx2 L3649 pTriplEx2 L3653 pBLUESCRIPT ™ sk(−) L3655 pBLUESCRIPT ™ sk(−) L3657 pBLUESCRIPT ™ sk(−) L3658 pTriplEx2 L3659 pBLUESCRIPT ™ L3661 pBLUESCRIPT ™ sk(−) L3665 kidney pDNR-LIB (CLONTECH ™) L3667 placenta pDNR-LIB (CLONTECH ™) L3684 breast puc18 L3705 colon puc18 L3713 colon puc18 L3722 placenta_normal puc18 L3729 placenta_normal puc18 L3744 head_neck puc18 L3750 head_neck puc18 L3783 testis_normal puc18 L3807 uterus_tumor puc18 L3808 uterus_tumor puc18 L3811 pBLUESCRIPT ™ sk(−) L3812 pBLUESCRIPT ™ sk(−) L3813 pTriplEx2 L3814 pTriplEx2 L3815 pTriplEx2 L3816 pME18SFL3 L3817 pME18SFL3 L3819 liver pDNR-LIB (CLONTECH ™) L3824 NT2 pME18SFL3 L3825 NT2 pME18SFL3 L3826 NT2 pUC19FL3 L3827 NT2 pME18SFL3 L3828 NT2 pME18SFL3 L3829 NT2 pME18SFL3 L3831 pME18SFL3 L3832 pME18SFL3 L3833 pME18SFL3 L3834 pME18SFL3 L3837 pME18SFL3 L3839 Y79 pME18SFL3 L3841 lung pOTB7 L3871 brain pOTB7 L3872 skin, pCMV-SPORT6 normal, 4 pooled sa L3904 brain pCMV-SPORT6 L3905 brain pCMV-SPORT6 L4501 pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L4537 thyroid pAMP10 L4556 tongue pCMV-SPORT6 L4560 uterus pCMV-SPORT6 L4669 ovary pCMV-SPORT6 L4747 brain pT7T3D-Pac (PHARMACIA ™) with a modified polylinker L5286 thyroid pAMP10 L5564 normal pAMP1 head/neck tissue L5565 brain pCMV-SPORT6 L5566 brain pCMV-SPORT6.ccdb L5568 head/neck pAMP1 L5569 nasopharynx pAMP10 L5574 nasopharynx pAMP10 L5575 brain pCMV-SPORT6 L5622 skin pCMV-SPORT6 L5623 skin pCMV-SPORT6

Description of Table 5

Table 5 provides a key to the OMIM reference identification numbers disclosed in Table 1B.1, column 9. OMIM reference identification numbers (Column 1) were derived from Online Mendelian Inheritance in Man (Online Mendelian Inheritance in Man, OMIM. McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University (Baltimore, Md.) and National Center for Biotechnology Information, National Library of Medicine, (Bethesda, Md.) 2000. Column 2 provides diseases associated with the cytologic band disclosed in Table 1B.1, column 8, as determined using the Morbid Map database.

TABLE 5 OMIM Reference Description 100710 Myasthenic syndrome, slow-channel congenital, 601462 101000 Meningioma, NF2-related, sporadic Schwannoma, sporadic 101000 Neurofibromatosis, type 2 101000 Neurolemmomatosis 101000 Malignant mesothelioma, sporadic 102200 Somatotrophinoma 102578 Leukemia, acute promyelocytic, PML/RARA type 102770 Myoadenylate deaminase deficiency 102772 [AMP deaminase deficiency, erythrocytic] 103050 Autism, succinylpurinemic 103050 Adenylosuccinase deficiency 103581 Albright hereditary osteodystrophy-2 103600 [Dysalbuminemic hyperthyroxinemia] 103600 [Dysalbuminemic hyperzincemia], 194470 103600 Analbuminemia 103850 Aldolase A deficiency 104150 [AFP deficiency, congenital] 104150 [Hereditary persistence of alpha-fetoprotein] 104500 Amelogenesis imperfecta-2, hypoplastic local type 104770 Amyloidosis, secondary, susceptibility to 106100 Angioedema, hereditary 106150 Hypertension, essential, susceptibility to 106150 Preeclampsia, susceptibility to 106165 Hypertension, essential, 145500 106180 Myocardial infarction, susceptibility to 106210 Peters anomaly 106210 Cataract, congenital, with late-onset corneal dystrophy 106210 Foveal hypoplasia, isolated, 136520 106210 Aniridia 107271 CD59 deficiency 107300 Antithrombin III deficiency 107670 Apolipoprotein A-II deficiency 107741 Hyperlipoproteinemia, type III 107776 Colton blood group, 110450 107777 Diabetes insipidus, nephrogenic, autosomal recessive, 222000 108725 Atherosclerosis, susceptibility to 108985 Atrophia areata 109270 Renal tubular acidosis, distal, 179800 109270 Spherocytosis, hereditary 109270 [Acanthocytosis, one form] 109270 [Elliptocytosis, Malaysian-Melanesian type] 109270 Hemolytic anemia due to band 3 defect 109400 Basal cell nevus syndrome 109560 Leukemia/lymphoma, B-cell, 3 109690 Asthma, nocturnal, susceptibility to 109690 Obesity, susceptibility to 109700 Hemodialysis-related amyloidosis 110100 Blepharophimosis, epicanthus inversus, and ptosis, type 1 110700 Vivax malaria, susceptibility to 113100 Brachydactyly, type C 113900 Heart block, progressive familial, type I 114550 Hepatocellular carcinoma 114835 Monocyte carboxyesterase deficiency 115500 Acatalasemia 115665 Cataract, congenital, Volkmann type 116800 Cataract, Marner type 116806 Colorectal cancer 116860 Cavernous angiomatous malformations 117700 [Hypoceruloplasminemia, hereditary] 117700 Hemosiderosis, systemic, due to aceruloplasminemia 118485 Polycystic ovary syndrome with hyperandrogenemia 118800 Choreoathetosis, familial paroxysmal 120070 Alport syndrome, autosomal recessive, 203780 120120 Epidermolysis bullosa dystrophica, dominant, 131750 120120 Epidermolysis bullosa dystrophica, recessive, 226600 120120 Epidermolysis bullosa, pretibial, 131850 120131 Alport syndrome, autosomal recessive, 203780 120131 Hematuria, familial benign 120140 Osteoarthrosis, precocious 120140 SED congenita 120140 SMED Strudwick type 120140 Stickler syndrome, type I 120140 Wagner syndrome, type II 120140 Achondrogenesis-hypochondrogenesis, type II 120140 Kniest dysplasia 120150 Osteogenesis imperfecta, 4 clinical forms, 166200, 166210, 259420, 166220 120150 Osteoporosis, idiopathic, 166710 120150 Ehlers-Danlos syndrome, type VIIA1, 130060 120215 Ehlers-Danlos syndrome, type I, 130000 120215 Ehlers-Danlos syndrome, type II, 130010 120220 Bethlem myopathy, 158810 120240 Bethlem myopathy, 158810 120260 Epiphyseal dysplasia, multiple, type 2, 600204 120435 Muir-Tone syndrome, 158320 120435 Colorectal cancer, hereditary, nonpolyposis, type 1 Ovarian cancer 120436 Muir-Tone family cancer syndrome, 158320 120436 Turcot syndrome with glioblastoma, 276300 120436 Colorectal cancer, hereditary nonpolyposis, type 2 120550 Clq deficiency, type A 120570 Clq deficiency, type B 120575 Clq deficiency, type C 120700 C3 deficiency 120950 C8 deficiency, type I 120960 C8 deficiency, type II 121011 Deafness, autosomal dominant 3, 601544 121011 Deafness, autosomal recessive 1, 220290 121050 Contractural arachnodactyly, congenital 121360 Myeloid leukemia, acute, M4Eo subtype 121800 Corneal dystrophy, crystalline, Schnyder 122720 Nicotine addiction, protection from 122720 Coumarin resistance, 122700 123000 Craniometaphyseal dysplasia 123101 Craniosynostosis, type 2 123270 [Creatine kinase, brain type, ectopic expression of] 123580 Cataract, congenital, autosomal dominant 123620 Cataract, cerulean, type 2, 601547 123660 Cataract, Coppock-like 123940 White sponge nevus, 193900 124030 Parkinsonism, susceptibility to 124030 Debrisoquine sensitivity 124200 Darier disease (keratosis follicularis) 125270 Porphyria, acute hepatic 125270 Lead poisoning, susceptibility to 125370 Dentatorubro-pallidoluysian atrophy 125660 Myopathy, desminopathic 125660 Cardiomyopathy 126090 Hyperphenylalaninemia due to pterin-4a-carbinolamine dehydratase deficiency, 264070 126337 Myxoid liposarcoma 126340 Xeroderma pigmentosum, group D, 278730 126391 DNA ligase I deficiency 126600 Drusen, radial, autosomal dominant 128100 Dystonia-1, torsion 129010 Neuropathy, congenital hypomyelinating, 1 129500 Ectodermal dysplasia, hidrotic 129900 EEC syndrome-1 130410 Glutaricaciduria, type IIB 130500 Elliptocytosis-1 131100 Multiple endocrine neoplasia I 131100 Prolactinoma, hyperparathyroidism, carcinoid syndrome 131100 Carcinoid tumor of lung 131210 Atherosclerosis, susceptibility to 131244 Hirschsprung disease-2, 600155 131400 Eosinophilia, familial 132700 Cylindromatosis 132800 Basal cell carcinoma 132800 Epithelioma, self-healing, squamous 1, Ferguson-Smith type 133171 [Erythrocytosis, familial], 133100 133200 Erythrokeratodermia variabilis 133530 Xeroderma pigmentosum, group G, 278780 133701 Exostoses, multiple, type 2 133780 Vitreoretinopathy, exudative, familial 134790 Hyperferritinemia-cataract syndrome, 600886 135300 Fibromatosis, gingival 135940 Ichthyosis vulgaris, 146700 136132 [Fish-odor syndrome], 602079 136350 Pfeiffer syndrome, 101600 136435 Ovarian dysgenesis, hypergonadotropic, with normal karyotype, 233300 136530 Male infertility, familial 136550 Macular dystrophy, North Carolina type 136836 Fucosyltransferase-6 deficiency 137350 Amyloidosis, Finnish type, 105120 138030 [Hyperproglucagonemia] 138040 Cortisol resistance 138079 Hyperinsulinism, familial, 602485 138079 MODY, type 2, 125851 138140 Glucose transport defect, blood-brain barrier 138160 Diabetes mellitus, noninsulin-dependent 138160 Fanconi-Bickel syndrome, 227810 138300 Hemolytic anemia due to glutathione reductase deficiency 138320 Hemolytic anemia due to glutathione peroxidase deficiency 138570 Non-insulin dependent diabetes mellitus, susceptibility to 138700 [Apolipoprotein H deficiency] 138981 Pulmonary alveolar proteinosis, 265120 139190 Gigantism due to GHRF hypersecretion 139190 Isolated growth hormone deficiency due to defect in GHRF 139191 Growth hormone deficient dwarfism 139250 Isolated growth hormone deficiency, Illig type with absent GH and Kowarski type with bioinactive GH 139350 Epidermolytic hyperkeratosis, 113800 139350 Keratoderma, palmoplantar, nonepidermolytic 140100 [Anhaptoglobinemia] 140100 [Hypohaptogloginemia] 141750 Alpha-thalassemia/mental retardation syndrome, type 1 141800 Methemoglobinemias, alpha- 141800 Thalassemias, alpha- 141800 Erythremias, alpha- 141800 Heinz body anemias, alpha- 141850 Thalassemia, alpha- 141850 Erythrocytosis 141850 Heinz body anemia 141850 Hemoglobin H disease 141850 Hypochromic microcytic anemia 142335 Hereditary persistence of fetal hemoglobin, heterocellular, Indian type 142600 Hemolytic anemia due to hexokinase deficiency 143890 Hypercholesterolemia, familial 144120 Hyperimmunoglobulin G1 syndrome 145001 Hyperparathyroidism-jaw tumor syndrome 145260 Pseudohypoaldosteronism, type II 145981 Hypocalciuric hypercalcemia, type II 146150 Hypomelanosis of Ito 146200 Hypoparathyroidism, familial 146760 [IgG receptor I, phagocytic, familial deficiency of] 146790 Lupus nephritis, susceptibility to 147020 Agammaglobulinemia, 601495 147050 Atopy 147110 IgG2 deficiency, selective 147141 Leukemia, acute lymphoblastic 147440 Growth retardation with deafness and mental retardation 147670 Rabson-Mendenhall syndrome 147670 Diabetes mellitus, insulin-resistant, with acanthosis nigricans 147670 Leprechaunism 147781 Atopy, susceptibility to 148040 Epidermolysis bullosa simplex, Koebner, Dowling-Meara, and Weber- Cockayne types, 131900, 131760, 131800 148041 Pachyonychia congenita, Jadassohn-Lewandowsky type, 167200 148043 Meesmann corneal dystrophy, 122100 148065 White sponge nevus, 193900 148070 Liver disease, susceptibility to, from hepatotoxins or viruses 148080 Epidermolytic hyperkeratosis, 113800 148370 Keratolytic winter erythema 148900 Klippel-Feil syndrome with laryngeal malformation 150200 [Placental lactogen deficiency] 150210 Lactoferrin-deficient neutrophils, 245480 150250 Larsen syndrome, autosomal dominant 150292 Epidermolysis bullosa, Herlitz junctional type, 226700 151385 Leukemia, acute myeloid 151390 Leukemia, acute T-cell 151410 Leukemia, chronic myeloid 151440 Leukemia, T-cell acute lymphoblastoid 151670 Hepatic lipase deficiency 152200 Coronary artery disease, susceptibility to 152427 Long QT syndrome-2 152445 Vohwinkel syndrome, 124500 152445 Erythrokeratoderma, progressive symmetric, 602036 152760 Hypogonadotropic hypogonadism due to GNRH deficiency, 227200 152780 Hypogonadism, hypergonadotropic 152780 Male pseudohermaphroditism due to defective LH 152790 Precocious puberty, male, 176410 152790 Leydig cell hypoplasia 153454 Ehlers-Danlos syndrome, type VI, 225400 153455 Cutis laxa, recessive, type I, 219100 153700 Macular dystrophy, vitelliform type 154275 Malignant hyperthermia susceptibility 2 154276 Malignant hyperthermia susceptibility 3 154400 Acrofacial dysostosis, Nager type 154545 Chronic infections, due to opsonin defect 154550 Carbohydrate-deficient glycoprotein syndrome, type Ib, 602579 156845 Tietz syndrome, 103500 156845 Waardenburg syndrome, type IIA, 193510 156845 Waardenburg syndrome/ocular albinism, digenic, 103470 156850 Cataract, congenital, with microphthalmia 157147 Abetalipoproteinemia, 200100 157170 Holoprosencephaly-2 157640 PEO with mitochondrial DNA deletions, type 1 157900 Moebius syndrome 158590 Spinal muscular atrophy-4 159000 Muscular dystrophy, limb-girdle, type 1A 159001 Muscular dystrophy, limb-girdle, type 1B 160760 Cardiomyopathy, familial hypertrophic, 1, 192600 160760 Central core disease, one form 160781 Cardiomyopathy, hypertrophic, mid-left ventricular chamber type 160900 Myotonic dystrophy 161015 Mitochondrial complex I deficiency, 252010 162150 Obestiy with impaired prohormone processing, 600955 162200 Neurofibromatosis, type 1 162200 Watson syndrome, 193520 162400 Neuropathy, hereditary sensory and autonomic, type 1 163729 Hypertension, pregnancy-induced 163950 Noonan syndrome-1 163950 Cardiofaciocutaneous syndrome, 115150 164009 Leukemia, acute promyelocytic, NUMA/RARA type 164500 Spinocerebellar ataxia-7 164731 Ovarian carcinoma, 167000 164770 Myeloid malignancy, predisposition to 164920 Piebaldism 164920 Mast cell leukemia 164920 Mastocytosis with associated hematologic disorder 164953 Liposarcoma 165240 Pallister-Hall syndrome, 146510 165240 Postaxial polydactyly type A1, 174200 165240 Greig cephalopolysyndactyly syndrome, 175700 165320 Hepatocellular carcinoma 167000 Ovarian cancer, serous 167410 Rhabdomyosarcoma, alveolar, 268220 168360 Paraneoplastic sensory neuropathy 168450 Hypoparathyroidism, autosomal dominant 168450 Hypoparathyroidism, autosomal recessive 168461 Multiple myeloma, 254250 168461 Parathyroid adenomatosis 1 168461 Centrocytic lymphoma 168468 Metaphyseal chondrodysplasia, Murk Jansen type, 156400 168500 Parietal foramina 169600 Hailey-Hailey disease 170500 Myotonia congenita, atypical acetazolamide-responsive 170500 Paramyotonia congenita, 168300 170500 Hyperkalemic periodic paralysis 170650 Periodontitis, juvenile 171190 Hypertension, essential, 145500 171650 Lysosomal acid phosphatase deficiency 171760 Hypophosphatasia, adult, 146300 171760 Hypophosphatasia, infantile, 241500 171860 Hemolytic anemia due to phosphofructokinase deficiency 172400 Hemolytic anemia due to glucosephosphate isomerase deficiency 172400 Hydrops fetalis, one form 172430 Enolase deficiency 172471 Glycogenosis, hepatic, autosomal 172490 Phosphorylase kinase deficiency of liver and muscle, 261750 173610 Platelet alpha/delta storage pool deficiency 173850 Polio, susceptibility to 173870 Xeroderma pigmentosum 173870 Fanconi anemia 173910 Polycystic kidney disease, adult, type II 174000 Medullary cystic kidney disease, AD 174900 Polyposis, juvenile intestinal 175100 Turcot syndrome, 276300 175100 Adenomatous polyposis coli 175100 Adenomatous polyposis coli, attenuated 175100 Colorectal cancer 175100 Desmoid disease, hereditary, 135290 175100 Gardner syndrome 176100 Porphyria cutanea tarda 176100 Porphyria, hepatoerythropoietic 176261 Jervell and Lange-Nielsen syndrome, 220400 176270 Prader-Willi syndrome 176450 Sacral agenesis-1 176830 Obesity, adrenal insufficiency, and red hair 176830 ACTH deficiency 176930 Dysprothrombinemia 176930 Hypoprothrombinemia 176960 Pituitary tumor, invasive 177400 Apnea, postanesthetic 178300 Ptosis, hereditary congenital, 1 178640 Pulmonary alveolar proteinosis, congenital, 265120 179095 Male infertility 179615 Reticulosis, familial histiocytic, 267700 179615 Severe combined immunodeficiency, B cell-negative, 601457 179616 Severe combined immunodeficiency, B cell-negative, 601457 179755 Renal cell carcinoma, papillary, 1 180071 Retinitis pigmentosa, autosomal recessive 180100 Retinitis pigmentosa-1 180104 Retinitis pigmentosa-9 180105 Retinitis pigmentosa-10 180380 Night blindness, congenital stationery, rhodopsin-related 180380 Retinitis pigmentosa, autosomal recessive 180380 Retinitis pigmentosa-4, autosomal dominant 180385 Leukemia, acute T-cell 180721 Retinitis pigmentosa, digenic 180840 Susceptibility to IDDM 180901 Malignant hyperthermia susceptibility 1, 145600 180901 Central core disease, 117000 181405 Scapuloperoneal spinal muscular atrophy, New England type 181430 Scapuloperoneal syndrome, myopathic type 181460 Schistosoma mansoni, susceptibility/resistance to 182138 Anxiety-related personality traits 182280 Small-cell cancer of lung 182290 Smith-Magenis syndrome 182380 Glucose/galactose malabsorption 182381 Renal glucosuria, 253100 182600 Spastic paraplegia-3A 182601 Spastic paraplegia-4 182860 Pyropoikilocytosis 182860 Spherocytosis, recessive 182860 Elliptocytosis-2 182900 Spherocytosis-2 185800 Symphalangism, proximal 186580 Arthrocutaneouveal granulomatosis 186855 Leukemia-2, T-cell acute lymphoblastic 186880 Leukemia/lymphoma, T-cell 186921 Leukemia, T-cell acute lymphoblastic 187040 Leukemia-1, T-cell acute lymphoblastic 188070 Bleeding disorder due to defective thromboxane A2 receptor 188450 Goiter, adolescent multinodular 188450 Goiter, nonendemic, simple 188450 Hypothyroidism, hereditary congenital 188826 Sorsby fundus dystrophy, 136900 189800 Preeclampsia/eclampsia 190040 Meningioma, SIS-related 190040 Dermatofibrosarcoma protuberans 190040 Giant-cell fibroblastoma 190195 Ichthyosiform erythroderma, congenital, 242100 190195 Ichthyosis, lamellar, autosomal recessive, 242300 190198 Leukemia, T-cell acute lymphoblastic 190300 Tremor, familial essential, 1 190605 Triphalangeal thumb-polysyndactyly syndrome 190685 Down syndrome 191044 Cardiomyopathy, familial hypertrophic 191092 Tuberous sclerosis-2 191100 Tuberous sclerosis-1 191181 Cervical carcinoma 191315 Insensitivity to pain, congenital, with anhidrosis, 256800 192090 Ovarian carcinoma 192090 Breast cancer, lobular 192090 Endometrial carcinoma 192090 Gastric cancer, familial, 137215 192974 Neonatal alloimmune thrombocytopenia 192974 Glycoprotein Ia deficiency 193235 Vitreoretinopathy, neovascular inflammatory 193500 Rhabdomyosarcoma, alveolar, 268220 193500 Waardenburg syndrome, type I 193500 Waardenburg syndrome, type III, 148820 193500 Craniofacial-deafness-hand syndrome, 122880 194070 Wilms tumor, type 1 194070 Denys-Drash syndrome 194070 Frasier syndrome, 136680 201460 Acyl-CoA dehydrogenase, long chain, deficiency of 201475 VLCAD deficiency 201810 3-beta-hydroxysteroid dehydrogenase, type II, deficiency 203300 Hermansky-Pudlak syndrome 203310 Ocular albinism, autosomal recessive 203500 Alkaptonuria 203740 Alpha-ketoglutarate dehydrogenase deficiency 205100 Amyotrophic lateral sclerosis, juvenile 205900 Anemia, Diamond-Blackfan 207750 Hyperlipoproteinemia, type Ib 208250 Jacobs syndrome 208400 Aspartylglucosaminuria 209901 Bardet-Biedl syndrome 1 212138 Carnitine-acylcarnitine translocase deficiency 215700 Citrullinemia 216550 Cohen syndrome 216900 Achromatopsia 217300 Cornea plana congenita, recessive 217800 Macular corneal dystrophy 218000 Andermann syndrome 218030 Apparent mineralocorticoid excess, hypertension due to 219800 Cystinosis, nephropathic 221770 Polycystic lipomembranous osteodysplasia with sclerosing leukencephalopathy 221820 Gliosis, familial progressive subcortical 222700 Lysinuric protein intolerance 222800 Hemolytic anemia due to bisphosphoglycerate mutase deficiency 222900 Sucrose intolerance 223360 Dopamine-beta-hydroxylase deficiency 223900 Dysautonomia, familial 224100 Congenital dyserythropoietic anemia II 225500 Ellis-van Creveld syndrome 227220 [Eye color, brown] 227645 Fanconi anemia, type C 229700 Fructose-bisphosphatase deficiency 229800 [Fructosuria] 230000 Fucosidosis 230400 Galactosemia 230800 Gaucher disease 230800 Gaucher disease with cardiovascular calcification 231550 Achalasia-addisonianism-alacrimia syndrome 231670 Glutaricaciduria, type I 231675 Glutaricaciduria, type IIC 231680 Glutaricaciduria, type IIA 232050 Propionicacidemia, type II or pccB type 232300 Glycogen storage disease II 232600 McArdle disease 232700 Glycogen storage disease VI 232800 Glycogen storage disease VII 233700 Chronic granulomatous disease due to deficiency of NCF-1 236100 Holoprosencephaly-1 236200 Homocystinuria, B6-responsive and nonresponsive types 236250 Homocystinuria due to MTHFR deficiency 236730 Urofacial syndrome 237300 Carbamoylphosphate synthetase I deficiency 238310 Hyperglycinemia, nonketotic, type II 238600 Chylomicronemia syndrome, familial 238600 Combined hyperlipemia, familial 238600 Hyperlipoproteinemia I 238600 Lipoprotein lipase deficiency 239100 Van Buchem disease 240300 Autoimmune polyglandular disease, type I 240400 Scurvy 245200 Krabbe disease 245349 Lacticacidemia due to PDX1 deficiency 245900 Norum disease 245900 Fish-eye disease 246450 HMG-CoA lyase deficiency 248510 Mannosidosis, beta- 248600 Maple syrup urine disease, type Ia 249000 Meckel syndrome 250250 Cartilage-hair hypoplasia 250790 Methemoglobinemia due to cytochrome b5 deficiency 250850 Hypermethioninemia, persistent, autosomal dominant, due to methionine adenosyltransferase I/III deficiency 251170 Mevalonicaciduria 251600 Microphthalmia, autosomal recessive 252500 Mucolipidosis II 252500 Mucolipidosis III 252900 Sanfilippo syndrome, type A 253250 Mulibrey nanism 253800 Walker-Warburg syndrome, 236670 253800 Fukuyama type congenital muscular dystrophy 255800 Schwartz-Jampel syndrome 256030 Nemaline myopathy-2 256540 Galactosialidosis 256700 Neuroblastoma 256731 Ceroid-lipofuscinosis, neuronal-5, variant late infantile 257200 Niemann-Pick disease, type A 257200 Niemann-Pick disease, type B 258501 3-methylglutaconicaciduria, type III 258900 Oroticaciduria 259700 Osteopetrosis, recessive 259770 Osteoporosis-pseudoglioma syndrome 259900 Hyperoxaluria, primary, type 1 261670 Myopathy due to phosphoglycerate mutase deficiency 262000 Bjornstad syndrome 266200 Anemia, hemolytic, due to PK deficiency 267750 Knobloch syndrome 268900 [Sarcosinemia] 269920 Salla disease 270100 Situs inversus viscerum 270200 Sjogren-Larsson syndrome 272750 GM2-gangliosidosis, AB variant 272800 Tay-Sachs disease 272800 [Hex A pseudodeficiency] 272800 GM2-gangliosidosis, juvenile, adult 276600 Tyrosinemia, type II 276700 Tyrosinemia, type I 276710 Tyrosinemia, type III 276900 Usher syndrome, type 1A 276901 Usher syndrome, type 2 276902 Usher syndrome, type 3 277700 Werner syndrome 277730 Wernicke-Korsakoff syndrome, susceptibility to 278700 Xeroderma pigmentosum, group A 300000 Opitz G syndrome, type I 300008 Nephrolithiasis, type I, 310468 300008 Proteinuria, low molecular weight, with hypercalciuric nephrocalcinosis 300008 Dent disease, 300009 300008 Hypophosphatemia, type III 300011 Menkes disease, 309400 300011 Occipital horn syndrome, 304150 300011 Cutis laxa, neonatal 300031 Mental retardation, X-linked, FRAXF type 300044 Wernicke-Korsakoff syndrome, susceptibility to 300046 Mental retardation, X-linked 23, nonspecific 300047 Mental retardation, X-linked 20 300048 Intestinal pseudoobstruction, neuronal, X-linked 300049 Nodular heterotopia, bilateral periventricular 300049 BPNH/MR syndrome 300055 Mental retardation with psychosis, pyramidal signs, and macroorchidism 300066 Deafness, X-linked 6, sensorineural 300071 Night blindness, congenital stationary, type 2 300075 Coffin-Lowry syndrome, 303600 300077 Mental retardation, X-linked 29 300100 Adrenoleukodystrophy 300100 Adrenomyeloneuropathy 300104 Mental retardation, X-linked nonspecific, 309541 300110 Night blindness, congenital stationary, X-linked incomplete, 300071 300123 Mental retardation with isolated growth hormone deficiency 300126 Dyskeratosis congenita-1, 305000 300127 Mental retardation, X-linked, 60 300310 Agammaglobulinemia, type 2, X-linked 300600 Ocular albinism, Forsius-Eriksson type 301000 Thrombocytopenia, X-linked, 313900 301000 Wiskott-Aldrich syndrome 301200 Amelogenesis imperfecta 301201 Amelogenesis imperfecta-3, hypoplastic type 301220 Partington syndrome II 301590 Anophthalmos-1 301830 Arthrogryposis, X-linked (spinal muscular atrophy, infantile, X-linked) 301835 Arts syndrome 301845 Bazex syndrome 302060 Noncompaction of left ventricular myocardium, isolated 302060 Barth syndrome 302060 Cardiomyopathy, X-linked dilated, 300069 302060 Endocardial fibroelastosis-2 302350 Nance-Horan syndrome 302801 Charcot-Marie-Tooth neuropathy, X-linked-2, recessive 302960 Chondrodysplasia punctata, X-linked dominant 303700 Colorblindness, blue monochromatic 303800 Colorblindness, deutan 303900 Colorblindness, protan 304050 Aicardi syndrome 304110 Craniofrontonasal dysplasia 304800 Diabetes insipidus, nephrogenic 305435 Heterocellular hereditary persistence of fetal hemoglobin, Swiss type 305450 FG syndrome 305900 Favism 305900 G6PD deficiency 305900 Hemolytic anemia due to G6PD deficiency 306000 Glycogenosis, X-linked hepatic, type I 306000 Glycogenosis, X-linked hepatic, type II 306100 Gonadal dysgenesis, XY female type 306700 Hemophilia A 306995 [Homosexuality, male] 307150 Hypertrichosis, congenital generalized 307800 Hypophosphatemia, hereditary 308310 Incontinentia pigmenti, familial 308800 Keratosis follicularis spinulosa decalvans 308840 Spastic paraplegia, 312900 308840 Hydrocephalus due to aqueductal stenosis, 307000 308840 MASA syndrome, 303350 309200 Manic-depressive illness, X-linked 309470 Mental retardation, X-linked, syndromic-3, with spastic diplegia 309500 Renpenning syndrome-1 309510 Mental retardation, X-linked, syndromic-1, with dystonic movements, ataxia, and seizures 309530 Mental retardation, X-linked 1, non-dysmorphic 309548 Mental retardation, X-linked, FRAXE type 309585 Mental retardation, X-linked, syndromic-6, with gynecomastia and obesity 309605 Mental retardation, X-linked, syndromic-4, with congenital contractures and low fingertip arches 309610 Mental retardation, X-linked, syndromic-2, with dysmorphism and cerebral atrophy 309620 Mental retardation-skeletal dysplasia 309850 Brunner syndrome 309900 Mucopolysaccharidosis II 310300 Emery-Dreifuss muscular dystrophy 310400 Myotubular myopathy, X-linked 310460 Myopia-1 310460 Bornholm eye disease 310490 Cowchock syndrome 311050 Optic atrophy, X-linked 311200 Oral-facial-digital syndrome 1 311300 Otopalatodigital syndrome, type I 311510 Waisman parkinsonism-mental retardation syndrome 311850 Phosphoribosyl pyrophosphate synthetase-related gout 312040 N syndrome, 310465 312060 Properdin deficiency, X-linked 312170 Pyruvate dehydrogenase deficiency 312700 Retinoschisis 313400 Spondyloepiphyseal dysplasia tarda 313700 Perineal hypospadias 313700 Prostate cancer 313700 Spinal and bulbar muscular atrophy of Kennedy, 313200 313700 Breast cancer, male, with Reifenstein syndrome 313700 Androgen insensitivity, several forms 314300 Goeminne TKCR syndrome 314400 Cardiac valvular dysplasia-1 314580 Wieacker-Wolff syndrome 600040 Colorectal cancer 600045 Xeroderma pigmentosum, group E, subtype 2 600065 Leukocyte adhesion deficiency, 116920 600079 Colon cancer 600101 Deafness, autosomal dominant 2 600119 Muscular dystrophy, Duchenne-like, type 2 600119 Adhalinopathy, primary 600138 Retinitis pigmentosa-11 600140 Rubenstein-Taybi syndrome, 180849 600143 Epilepsy, progressive, with mental retardation 600163 Long QT syndrome-3 600173 SCID, autosomal recessive, T-negative/B-positive type 600175 Spinal muscular atrophy, congenital nonprogressive, of lower limbs 600194 Ichthyosis bullosa of Siemens, 146800 600223 Spinocerebellar ataxia-4 600231 Palmoplantar keratoderma, Bothnia type 600234 HMG-CoA synthease-2 deficiency 600243 Temperature-sensitive apoptosis 600259 Turcot syndrome with glioblastoma, 276300 600259 Colorectal cancer, hereditary nonpolyposis, type 4 600266 Resistance/susceptibility to TB, etc. 600273 Polycystic kidney disease, infantile severe, with tuberous sclerosis 600276 Cerebral arteriopathy with subcortical infarcts and leukoencephalopathy, 125310 600281 Non-insulin-dependent diabetes mellitus, 125853 600281 MODY, type 1, 125850 600309 Atrioventricular canal defect-1 600310 Pseudoachondroplasia, 177170 600310 Epiphyseal dysplasia, multiple 1, 132400 600319 Diabetes mellitus, insulin-dependent, 4 600320 Insulin-dependent diabetes mellitus-5 600332 Rippling muscle disease-1 600374 Bardet-Biedl syndrome 4 600510 Pigment dispersion syndrome 600512 Epilepsy, partial 600525 Trichodontoosseous syndrome, 190320 600528 CPT deficiency, hepatic, type I, 255120 600536 Myopathy, congenital 600584 Atrial septal defect with atrioventricular conduction defects, 108900 600593 Craniosynostosis, Adelaide type 600617 Lipoid adrenal hyperplasia, 201710 600623 Prostate cancer, 176807 600631 Enuresis, nocturnal, 1 600650 Myopathy due to CPT II deficiency, 255110 600650 CPT deficiency, hepatic, type II, 600649 600652 Deafness, autosomal dominant 4 600698 Salivary adenoma 600698 Uterine leiomyoma 600698 Lipoma 600698 Lipomatosis, mutiple, 151900 600722 Ceroid lipofuscinosis, neuronal, variant juvenile type, with granular osmiophilic deposits 600722 Ceroid lipofuscinosis, neuronal-1, infantile, 256730 600725 Holoprosencephaly-3, 142945 600757 Orofacial cleft-3 600759 Alzheimer disease-4 600792 Deafness, autosomal recessive 5 600807 Bronchial asthma 600808 Enuresis, nocturnal, 2 600811 Xeroderma pigmentosum, group E, DDB-negative subtype, 278740 600850 Schizophrenia disorder-4 600852 Retinitis pigmentosa-17 600881 Cataract, congenital, zonular, with sutural opacities 600882 Charcot-Marie-Tooth neuropathy-2B 600883 Diabetes mellitus, insulin-dependent, 8 600897 Cataract, zonular pulverulent-1, 116200 600900 Muscular dystrophy, limb-girdle, type 2E 600918 Cystinuria, type III 600956 Persistent Mullerian duct syndrome, type II, 261550 600957 Persistent Mullerian duct syndrome, type I, 261550 600958 Cardiomyopathy, familial hypertrophic, 4, 115197 600968 Gitelman syndrome, 263800 600971 Deafness, autosomal recessive 6 600975 Glaucoma 3, primary infantile, B 600995 Nephrotic syndrome, idiopathic, steroid-resistant 600996 Arrhythmogenic right ventricular dysplasia-2 601002 5-oxoprolinuria, 266130 601002 Hemolytic anemia due to glutathione synthetase deficiency, 231900 601072 Deafness, autosomal recessive 8 601097 Neuropathy, recurrent, with pressure palsies, 162500 601097 Charcot-Marie-Tooth neuropathy-1A, 118220 601097 Dejerine-Sottas disease, PMP22 related, 145900 601105 Pycnodysostosis, 265800 601145 Epilepsy, progressive myoclonic 1, 254800 601146 Brachydactyly, type C, 113100 601146 Acromesomelic dysplasia, Hunter-Thompson type, 201250 601146 Chondrodysplasia, Grebe type, 200700 601199 Neonatal hyperparathyroidism, 239200 601199 Hypocalcemia, autosomal dominant, 601198 601199 Hypocalciuric hypercalcemia, type I, 145980 601226 Progressive external ophthalmoplegia, type 2 601238 Cerebellar ataxia, Cayman type 601277 Ichthyosis, lamellar, type 2 601284 Hereditary hemorrhagic telangiectasia-2, 600376 601295 Bile acid malabsorption, primary 601309 Basal cell carcinoma, sporadic 601309 Basal cell nevus syndrome, 109400 601313 Polycystic kidney disease, adult type I, 173900 601369 Deafness, autosomal dominant 9 601385 Prostate cancer 601386 Deafness, autosomal recessive 12 601399 Platelet disorder, familial, with associated myeloid malignancy 601402 Leukemia, myeloid, acute 601412 Deafness, autosomal dominant 7 601414 Retinitis pigmentosa-18 601458 Inflammatory bowel disease-2 601471 Moebius syndrome-2 601472 Charcot-Marie-Tooth neuropathy-2D 601493 Cardiomyopathy, dilated 1C 601517 Spinocerebellar ataxia-2, 183090 601518 Prostate cancer, hereditary, 1, 176807 601596 Charcot-Marie-Tooth neuropathy, demyelinating 601604 Mycobacterial and salmonella infections, susceptibility to 601623 Angelman syndrome 601649 Blepharophimosis, epicanthus inversus, and ptosis, type 2 601652 Glaucoma 1A, primary open angle, juvenile-onset, 137750 601669 Hirschsprung disease, one form 601682 Glaucoma 1C, primary open angle 601691 Retinitis pigmentosa-19, 601718 601691 Stargardt disease-1, 248200 601691 Cone-rod dystrophy 3 601691 Fundus flavimaculatus with macular dystrophy, 248200 601692 Reis-Bucklers corneal dystrophy 601692 Corneal dystrophy, Avellino type 601692 Corneal dystrophy, Groenouw type I, 121900 601692 Corneal dystrophy, lattice type I, 122200 601718 Retinitis pigmentosa-19 601744 Systemic lupus erythematosus, susceptibility to, 1 601769 Osteoporosis, involutional 601769 Rickets, vitamin D-resistant, 277440 601771 Glaucoma 3A, primary infantile, 231300 601780 Ceroid-lipofuscinosis, neuronal-6, variant late infantile 601785 Carbohydrate-deficient glycoprotein syndrome, type I, 212065 601800 [Hair color, brown] 601843 Hypothyroidism, congenital, 274400 601844 Pseudohypoaldosteronism type II 601846 Muscular dystrophy with rimmed vacuoles 601850 Retinitis pigmentosa-deafness syndrome 601863 Bare lymphocyte syndrome, complementation group C 601884 [High bone mass] 601885 Cataract, zonular pulverulent-2 601889 Lymphoma, diffuse large cell 601928 Monilethrix, 158000 601954 Muscular dystrophy, limb-girdle, type 2G 601975 Ectodermal dysplasia/skin fragility syndrome 602025 Obesity/hyperinsulinism, susceptibility to 602085 Postaxial polydactyly, type A2 602086 Arrhythmogenic right ventricular dysplasia-3 602088 Nephronophthisis, infantile 602089 Hemangioma, capillary, hereditary 602092 Deafness, autosomal recessive 18 602094 Lipodystrophy, familial partial 602116 Glioma 602117 Prader-Willi syndrome 602121 Deafness, autosomal dominant nonsyndromic sensorineural, 1, 124900 602134 Tremor, familial essential, 2 602136 Refsum disease, infantile, 266510 602136 Zellweger syndrome-1, 214100 602136 Adrenoleukodystrophy, neonatal, 202370 602153 Monilethrix, 158000 602216 Peutz-Jeghers syndrome, 175200 602221 Stem-cell leukemia/lymphoma syndrome 602225 Cone-rod retinal dystrophy-2, 120970 602225 Leber congenital amaurosis, type III 602279 Oculopharyngeal muscular dystorphy, 164300 602279 Oculopharyngeal muscular dystrophy, autosomal recessive, 257950 602363 Ellis-van Creveld-like syndrome 602403 Alzheimer disease, susceptibility to 602447 Coronary artery disease, susceptibility to 602460 Deafness, autosomal dominant 15, 602459 602477 Febrile convulsions, familial, 2 602491 Hyperlipidemia, familial combined, 1 602544 Parkinson disease, juvenile, type 2, 600116 602568 Homocystinuria-megaloblastic anemia, cbl E type, 236270 602629 Dystonia-6, torsion 602666 Deafness, autosomal recessive 3, 600316 602716 Nephrosis-1, congenital, Finnish type, 256300 602772 Retinitis pitmentosa-24 602782 Faisalabad histiocytosis

Mature Polypeptides

The present invention also encompasses mature forms of a polypeptide having the amino acid sequence of SEQ ID NO:Y and/or the amino acid sequence encoded by the cDNA in a deposited clone. Polynucleotides encoding the mature forms (such as, for example, the polynucleotide sequence in SEQ ID NO:X and/or the polynucleotide sequence contained in the cDNA of a deposited clone) are also encompassed by the invention. Moreover, fragments or variants of these polypeptides (such as, fragments as described herein, polypeptides at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to these polypeptides, or polypeptides encoded by a polynucleotide that hybridizes under stringent conditions to the complementary strand of the polynucleotide encoding these polypeptides) are also encompassed by the invention. In preferred embodiments, these fragments or variants retain one or more functional activities of the full-length or mature form of the polypeptide (e.g., biological activity (such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune disorders), antigenicity (ability to bind, or compete with a polypeptide of the invention for binding, to an anti-polypeptide of the invention antibody), immunogenicity (ability to generate antibody which binds to a specific polypeptide of the invention), ability to form multimers with polypeptides of the invention, and ability to bind to a receptor or ligand for a polypeptide of the invention). Antibodies that bind the polypeptides of the invention, and polynucleotides encoding these polypeptides are also encompassed by the invention.

According to the signal hypothesis, proteins secreted by mammalian cells have a signal or secretary leader sequence that is cleaved from the mature protein once export of the growing protein chain across the rough endoplasmic reticulum has been initiated. Most mammalian cells and even insect cells cleave secreted proteins with the same specificity. However, in some cases, cleavage of a secreted protein is not entirely uniform, which results in two or more mature species of the protein. Further, it has long been known that cleavage specificity of a secreted protein is ultimately determined by the primary structure of the complete protein, that is, it is inherent in the amino acid sequence of the polypeptide.

Methods for predicting whether a protein has a signal sequence, as well as the cleavage point for that sequence, are available. For instance, the method of McGeoch, Virus Res. 3:271-286 (1985), uses the information from a short N-terminal charged region and a subsequent uncharged region of the complete (uncleaved) protein. The method of von Heinje, Nucleic Acids Res. 14:4683-4690 (1986) uses the information from the residues surrounding the cleavage site, typically residues −13 to +2, where +1 indicates the amino terminus of the secreted protein. The accuracy of predicting the cleavage points of known mammalian secretory proteins for each of these methods is in the range of 75-80%. (von Heinje, supra.) However, the two methods do not always produce the same predicted cleavage point(s) for a given protein.

In the present case, the deduced amino acid sequence of the secreted polypeptide was analyzed by a computer program called SignalP (Henrik Nielsen et al., Protein Engineering 10:1-6 (1997)), which predicts the cellular location of a protein based on the amino acid sequence. As part of this computational prediction of localization, the methods of McGeoch and von Heinje are incorporated. The analysis of the amino acid sequences of the secreted proteins described herein by this program provided the results shown in Table 1A.

In specific embodiments, polypeptides of the invention comprise, or alternatively consist of, the predicted mature form of the polypeptide as delineated in columns 14 and 15 of Table 1A. Moreover, fragments or variants of these polypeptides (such as, fragments as described herein, polypeptides at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to these polypeptides, or polypeptides encoded by a polynucleotide that hybridizes under stringent conditions to the complementary strand of the polynucleotide encoding these polypeptides) are also encompassed by the invention. In preferred embodiments, these fragments or variants retain one or more functional activities of the full-length or mature form of the polypeptide (e.g., biological activity (such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune disorders), antigenicity (ability to bind, or compete with a polypeptide of the invention for binding, to an anti-polypeptide of the invention antibody), immunogenicity (ability to generate antibody which binds to a specific polypeptide of the invention), ability to form multimers with polypeptides of the invention, and ability to bind to a receptor or ligand for a polypeptide of the invention). Antibodies that bind the polypeptides of the invention, and polynucleotides encoding these polypeptides are also encompassed by the invention.

Polynucleotides encoding proteins comprising, or consisting of, the predicted mature form of polypeptides of the invention (e.g., polynucleotides having the sequence of SEQ ID NO: X (Table 1A, column 4), the sequence delineated in columns 7 and 8 of Table 1A, and a sequence encoding the mature polypeptide delineated in columns 14 and 15 of Table 1A (e.g., the sequence of SEQ ID NO:X encoding the mature polypeptide delineated in columns 14 and 15 of Table 1)) are also encompassed by the invention, as are fragments or variants of these polynucleotides (such as, fragments as described herein, polynucleotides at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to these polynucleotides, and nucleic acids which hybridizes under stringent conditions to the complementary strand of the polynucleotide).

As one of ordinary skill would appreciate, however, cleavage sites sometimes vary from organism to organism and cannot be predicted with absolute certainty. Accordingly, the present invention provides secreted polypeptides having a sequence shown in SEQ ID NO:Y which have an N-terminus beginning within 15 residues of the predicted cleavage point (i.e., having 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, or 15 more or less contiguous residues of SEQ ID NO:Y at the N-terminus when compared to the predicted mature form of the polypeptide (e.g., the mature polypeptide delineated in columns 14 and 15 of Table 1). Similarly, it is also recognized that in some cases, cleavage of the signal sequence from a secreted protein is not entirely uniform, resulting in more than one secreted species. These polypeptides, and the polynucleotides encoding such polypeptides, are contemplated by the present invention.

Moreover, the signal sequence identified by the above analysis may not necessarily predict the naturally occurring signal sequence. For example, the naturally occurring signal sequence may be further upstream from the predicted signal sequence. However, it is likely that the predicted signal sequence will be capable of directing the secreted protein to the ER. Nonetheless, the present invention provides the mature protein produced by expression of the polynucleotide sequence of SEQ ID NO:X and/or the polynucleotide sequence contained in the cDNA of a deposited clone, in a mammalian cell (e.g., COS cells, as described below). These polypeptides, and the polynucleotides encoding such polypeptides, are contemplated by the present invention.

Polynucleotide and Polypeptide Variants

The present invention is also directed to variants of the polynucleotide sequence disclosed in SEQ ID NO:X or the complementary strand thereto, nucleotide sequences encoding the polypeptide of SEQ ID NO:Y, the nucleotide sequence of SEQ ID NO:X that encodes the polypeptide sequence as defined in columns 13 and 14 of Table 1A, nucleotide sequences encoding the polypeptide sequence as defined in columns 13 and 14 of Table 1A, the nucleotide sequence of SEQ ID NO:X encoding the polypeptide sequence as defined in Table 1B, nucleotide sequences encoding the polypeptide as defined in Table 1B, the nucleotide sequence as defined in columns 8 and 9 of Table 2, nucleotide sequences encoding the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2, the nucleotide sequence as defined in column 6 of Table 1C, nucleotide sequences encoding the polypeptide encoded by the nucleotide sequence as defined in column 6 of Table 1C, the cDNA sequence contained in ATCC™ Deposit No:Z, nucleotide sequences encoding the polypeptide encoded by the cDNA sequence contained in ATCC™ Deposit No:Z, and/or nucleotide sequences encoding a mature (secreted) polypeptide encoded by the cDNA sequence contained in ATCC™ Deposit No:Z.

The present invention also encompasses variants of the polypeptide sequence disclosed in SEQ ID NO:Y, the polypeptide as defined in columns 13 and 14 of Table 1A, the polypeptide sequence as defined in Table 1B, a polypeptide sequence encoded by the polynucleotide sequence in SEQ ID NO:X, a polypeptide sequence encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2, a polypeptide sequence encoded by the nucleotide sequence as defined in column 6 of Table 1C, a polypeptide sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X, the polypeptide sequence encoded by the cDNA sequence contained in ATCC™ Deposit No:Z and/or a mature (secreted) polypeptide encoded by the cDNA sequence contained in ATCC™ Deposit No:Z.

“Variant” refers to a polynucleotide or polypeptide differing from the polynucleotide or polypeptide of the present invention, but retaining essential properties thereof. Generally, variants are overall closely similar, and, in many regions, identical to the polynucleotide or polypeptide of the present invention.

Thus, one aspect of the invention provides an isolated nucleic acid molecule comprising, or alternatively consisting of, a polynucleotide having a nucleotide sequence selected from the group consisting of: (a) a nucleotide sequence described in SEQ ID NO:X or contained in the cDNA sequence of ATCC™ Deposit No:Z; (b) a nucleotide sequence in SEQ ID NO:X or the cDNA in ATCC™ Deposit No:Z which encodes the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z; (c) a nucleotide sequence in SEQ ID NO:X or the cDNA in ATCC™ Deposit No:Z which encodes a mature polypeptide (i.e., a secreted polypeptide (e.g., as delineated in columns 14 and 15 of Table 1A)); (d) a nucleotide sequence in SEQ ID NO:X or the cDNA sequence of ATCC™ Deposit No:Z, which encodes a biologically active fragment of a polypeptide; (e) a nucleotide sequence in SEQ ID NO:X or the cDNA sequence of ATCC™ Deposit No:Z, which encodes an antigenic fragment of a polypeptide; (f) a nucleotide sequence encoding a polypeptide comprising the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z; (g) a nucleotide sequence encoding a mature polypeptide of the amino acid sequence of SEQ ID NO:Y (i.e., a secreted polypeptide (e.g., as delineated in columns 14 and 15 of Table 1A)) or a mature polypeptide of the amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z; (h) a nucleotide sequence encoding a biologically active fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z; (i) a nucleotide sequence encoding an antigenic fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z; and (j) a nucleotide sequence complementary to any of the nucleotide sequences in (a), (b), (c), (d), (e), (f), (g), (h), or (i) above.

The present invention is also directed to nucleic acid molecules which comprise, or alternatively consist of, a nucleotide sequence which is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100%, identical to, for example, any of the nucleotide sequences in (a), (b), (c), (d), (e), (f), (g), (h), (i), or (j) above, the nucleotide coding sequence in SEQ ID NO:X or the complementary strand thereto, the nucleotide coding sequence of the cDNA contained in ATCC™ Deposit No:Z or the complementary strand thereto, a nucleotide sequence encoding the polypeptide of SEQ ID NO:Y, a nucleotide sequence encoding a polypeptide sequence encoded by the nucleotide sequence in SEQ ID NO:X, a polypeptide sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X, a nucleotide sequence encoding the polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z, the nucleotide coding sequence in SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto, a nucleotide sequence encoding the polypeptide encoded by the nucleotide sequence in SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto, the nucleotide coding sequence in SEQ ID NO:B as defined in column 6 of Table 1C or the complementary strand thereto, a nucleotide sequence encoding the polypeptide encoded by the nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1C or the complementary strand thereto, the nucleotide sequence in SEQ ID NO:X encoding the polypeptide sequence as defined in Table 1B or the complementary strand thereto, nucleotide sequences encoding the polypeptide as defined in Table 1B or the complementary strand thereto, and/or polynucleotide fragments of any of these nucleic acid molecules (e.g., those fragments described herein). Polynucleotides which hybridize to the complement of these nucleic acid molecules under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention, as are polypeptides encoded by these polynucleotides and nucleic acids.

In a preferred embodiment, the invention encompasses nucleic acid molecules which comprise, or alternatively, consist of a polynucleotide which hybridizes under stringent hybridization conditions, or alternatively, under lower stringency conditions, to a polynucleotide in (a), (b), (c), (d), (e), (f), (g), (h), or (i), above, as are polypeptides encoded by these polynucleotides. In another preferred embodiment, polynucleotides which hybridize to the complement of these nucleic acid molecules under stringent hybridization conditions, or alternatively, under lower stringency conditions, are also encompassed by the invention, as are polypeptides encoded by these polynucleotides.

In another embodiment, the invention provides a purified protein comprising, or alternatively consisting of, a polypeptide having an amino acid sequence selected from the group consisting of: (a) the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z; (b) the amino acid sequence of a mature (secreted) form of a polypeptide having the amino acid sequence of SEQ ID NO:Y (e.g., as delineated in columns 14 and 15 of Table 1A) or a mature form of the amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z mature; (c) the amino acid sequence of a biologically active fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z; and (d) the amino acid sequence of an antigenic fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in ATCC™ Deposit No:Z.

The present invention is also directed to proteins which comprise, or alternatively consist of, an amino acid sequence which is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100%, identical to, for example, any of the amino acid sequences in (a), (b), (c), or (d), above, the amino acid sequence shown in SEQ ID NO:Y, the amino acid sequence encoded by the cDNA contained in ATCC™ Deposit No:Z, the amino acid sequence of the polypeptide encoded by the nucleotide sequence in SEQ ID NO:X as defined in columns 8 and 9 of Table 2, the amino acid sequence of the polypeptide encoded by the nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1C, the amino acid sequence as defined in Table 1B, an amino acid sequence encoded by the nucleotide sequence in SEQ ID NO:X, and an amino acid sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X. Fragments of these polypeptides are also provided (e.g., those fragments described herein). Further proteins encoded by polynucleotides which hybridize to the complement of the nucleic acid molecules encoding these amino acid sequences under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention, as are the polynucleotides encoding these proteins.

By a nucleic acid having a nucleotide sequence at least, for example, 95% “identical” to a reference nucleotide sequence of the present invention, it is intended that the nucleotide sequence of the nucleic acid is identical to the reference sequence except that the nucleotide sequence may include up to five point mutations per each 100 nucleotides of the reference nucleotide sequence encoding the polypeptide. In other words, to obtain a nucleic acid having a nucleotide sequence at least 95% identical to a reference nucleotide sequence, up to 5% of the nucleotides in the reference sequence may be deleted or substituted with another nucleotide, or a number of nucleotides up to 5% of the total nucleotides in the reference sequence may be inserted into the reference sequence. The query sequence may be an entire sequence referred to in Table 1B or 2 as the ORF (open reading frame), or any fragment specified as described herein.

As a practical matter, whether any particular nucleic acid molecule or polypeptide is at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to a nucleotide sequence of the present invention can be determined conventionally using known computer programs. A preferred method for determining the best overall match between a query sequence (a sequence of the present invention) and a subject sequence, also referred to as a global sequence alignment, can be determined using the FASTDB computer program based on the algorithm of Brutlag et al. (Comp. App. Biosci. 6:237-245 (1990)). In a sequence alignment the query and subject sequences are both DNA sequences. An RNA sequence can be compared by converting U's to T's. The result of said global sequence alignment is expressed as percent identity. Preferred parameters used in a FASTDB alignment of DNA sequences to calculate percent identity are: Matrix=Unitary, k-tuple=4, Mismatch Penalty=1, Joining Penalty=30, Randomization Group Length=0, Cutoff Score=1, Gap Penalty=5, Gap Size Penalty 0.05, Window Size=500 or the length of the subject nucleotide sequence, whichever is shorter.

If the subject sequence is shorter than the query sequence because of 5′ or 3′ deletions, not because of internal deletions, a manual correction must be made to the results. This is because the FASTDB program does not account for 5′ and 3′ truncations of the subject sequence when calculating percent identity. For subject sequences truncated at the 5′ or 3′ ends, relative to the query sequence, the percent identity is corrected by calculating the number of bases of the query sequence that are 5′ and 3′ of the subject sequence, which are not matched/aligned, as a percent of the total bases of the query sequence. Whether a nucleotide is matched/aligned is determined by results of the FASTDB sequence alignment. This percentage is then subtracted from the percent identity, calculated by the above FASTDB program using the specified parameters, to arrive at a final percent identity score. This corrected score is what is used for the purposes of the present invention. Only bases outside the 5′ and 3′ bases of the subject sequence, as displayed by the FASTDB alignment, which are not matched/aligned with the query sequence, are calculated for the purposes of manually adjusting the percent identity score.

For example, a 90 base subject sequence is aligned to a 100 base query sequence to determine percent identity. The deletions occur at the 5′ end of the subject sequence and therefore, the FASTDB alignment does not show a matched/alignment of the first 10 bases at 5′ end. The 10 unpaired bases represent 10% of the sequence (number of bases at the 5′ and 3′ ends not matched/total number of bases in the query sequence) so 10% is subtracted from the percent identity score calculated by the FASTDB program. If the remaining 90 bases were perfectly matched the final percent identity would be 90%. In another example, a 90 base subject sequence is compared with a 100 base query sequence. This time the deletions are internal deletions so that there are no bases on the 5′ or 3′ of the subject sequence which are not matched/aligned with the query. In this case the percent identity calculated by FASTDB is not manually corrected. Once again, only bases 5′ and 3′ of the subject sequence which are not matched/aligned with the query sequence are manually corrected for. No other manual corrections are to be made for the purposes of the present invention.

By a polypeptide having an amino acid sequence at least, for example, 95% “identical” to a query amino acid sequence of the present invention, it is intended that the amino acid sequence of the subject polypeptide is identical to the query sequence except that the subject polypeptide sequence may include up to five amino acid alterations per each 100 amino acids of the query amino acid sequence. In other words, to obtain a polypeptide having an amino acid sequence at least 95% identical to a query amino acid sequence, up to 5% of the amino acid residues in the subject sequence may be inserted, deleted, (indels) or substituted with another amino acid. These alterations of the reference sequence may occur at the amino or carboxy terminal positions of the reference amino acid sequence or anywhere between those terminal positions, interspersed either individually among residues in the reference sequence or in one or more contiguous groups within the reference sequence.

As a practical matter, whether any particular polypeptide is at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to, for instance, the amino acid sequence of a polypeptide referred to in Table 1A (e.g., the amino acid sequence delineated in columns 14 and 15) or a fragment thereof, Table 1B (e.g., the amino acid sequence identified in column 6) or a fragment thereof, Table 2 (e.g., the amino acid sequence of the polypeptide encoded by the polynucleotide sequence defined in columns 8 and 9 of Table 2) or a fragment thereof, the amino acid sequence of the polypeptide encoded by the polynucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1C or a fragment thereof, the amino acid sequence of the polypeptide encoded by the nucleotide sequence in SEQ ID NO:X or a fragment thereof, or the amino acid sequence of the polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z, or a fragment thereof, the amino acid sequence of a mature (secreted) polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z, or a fragment thereof, can be determined conventionally using known computer programs. A preferred method for determining the best overall match between a query sequence (a sequence of the present invention) and a subject sequence, also referred to as a global sequence alignment, can be determined using the FASTDB computer program based on the algorithm of Brutlag et al. (Comp. App. Biosci. 6:237-245 (1990)). In a sequence alignment the query and subject sequences are either both nucleotide sequences or both amino acid sequences. The result of said global sequence alignment is expressed as percent identity. Preferred parameters used in a FASTDB amino acid alignment are: Matrix=PAM 0, k-tuple=2, Mismatch Penalty=1, Joining Penalty=20, Randomization Group Length=0, Cutoff Score=1, Window Size=sequence length, Gap Penalty=5, Gap Size Penalty=0.05, Window Size=500 or the length of the subject amino acid sequence, whichever is shorter.

If the subject sequence is shorter than the query sequence due to N- or C-terminal deletions, not because of internal deletions, a manual correction must be made to the results. This is because the FASTDB program does not account for N- and C-terminal truncations of the subject sequence when calculating global percent identity. For subject sequences truncated at the N- and C-termini, relative to the query sequence, the percent identity is corrected by calculating the number of residues of the query sequence that are N- and C-terminal of the subject sequence, which are not matched/aligned with a corresponding subject residue, as a percent of the total bases of the query sequence. Whether a residue is matched/aligned is determined by results of the FASTDB sequence alignment. This percentage is then subtracted from the percent identity, calculated by the above FASTDB program using the specified parameters, to arrive at a final percent identity score. This final percent identity score is what is used for the purposes of the present invention. Only residues to the N- and C-termini of the subject sequence, which are not matched/aligned with the query sequence, are considered for the purposes of manually adjusting the percent identity score. That is, only query residue positions outside the farthest N- and C-terminal residues of the subject sequence.

For example, a 90 amino acid residue subject sequence is aligned with a 100 residue query sequence to determine percent identity. The deletion occurs at the N-terminus of the subject sequence and therefore, the FASTDB alignment does not show a matching/alignment of the first 10 residues at the N-terminus. The 10 unpaired residues represent 10% of the sequence (number of residues at the N- and C-termini not matched/total number of residues in the query sequence) so 10% is subtracted from the percent identity score calculated by the FASTDB program. If the remaining 90 residues were perfectly matched the final percent identity would be 90%. In another example, a 90 residue subject sequence is compared with a 100 residue query sequence. This time the deletions are internal deletions so there are no residues at the N- or C-termini of the subject sequence which are not matched/aligned with the query. In this case the percent identity calculated by FASTDB is not manually corrected. Once again, only residue positions outside the N- and C-terminal ends of the subject sequence, as displayed in the FASTDB alignment, which are not matched/aligned with the query sequence are manually corrected for. No other manual corrections are to made for the purposes of the present invention.

The polynucleotide variants of the invention may contain alterations in the coding regions, non-coding regions, or both. Especially preferred are polynucleotide variants containing alterations which produce silent substitutions, additions, or deletions, but do not alter the properties or activities of the encoded polypeptide. Nucleotide variants produced by silent substitutions due to the degeneracy of the genetic code are preferred. Moreover, polypeptide variants in which less than 50, less than 40, less than 30, less than 20, less than 10, or 5-50, 5-25, 5-10, 1-5, or 1-2 amino acids are substituted, deleted, or added in any combination are also preferred. Polynucleotide variants can be produced for a variety of reasons, e.g., to optimize codon expression for a particular host (change codons in the human mRNA to those preferred by a bacterial host such as E. coli).

Naturally occurring variants are called “allelic variants,” and refer to one of several alternate forms of a gene occupying a given locus on a chromosome of an organism. (Genes II, Lewin, B., ed., John Wiley & Sons, New York (1985)). These allelic variants can vary at either the polynucleotide and/or polypeptide level and are included in the present invention. Alternatively, non-naturally occurring variants may be produced by mutagenesis techniques or by direct synthesis.

Using known methods of protein engineering and recombinant DNA technology, variants may be generated to improve or alter the characteristics of the polypeptides of the present invention. For instance, one or more amino acids can be deleted from the N-terminus or C-terminus of the polypeptide of the present invention without substantial loss of biological function. As an example, Ron et al. (J. Biol. Chem. 268: 2984-2988 (1993)) reported variant KGF proteins having heparin binding activity even after deleting 3, 8, or 27 amino-terminal amino acid residues. Similarly, Interferon gamma exhibited up to ten times higher activity after deleting 8-10 amino acid residues from the carboxy terminus of this protein. (Dobeli et al., J. Biotechnology 7:199-216 (1988).)

Moreover, ample evidence demonstrates that variants often retain a biological activity similar to that of the naturally occurring protein. For example, Gayle and coworkers (J. Biol. Chem. 268:22105-22111 (1993)) conducted extensive mutational analysis of human cytokine IL-1a. They used random mutagenesis to generate over 3,500 individual IL-1a mutants that averaged 2.5 amino acid changes per variant over the entire length of the molecule. Multiple mutations were examined at every possible amino acid position. The investigators found that “[m]ost of the molecule could be altered with little effect on either [binding or biological activity].” In fact, only 23 unique amino acid sequences, out of more than 3,500 nucleotide sequences examined, produced a protein that significantly differed in activity from wild-type.

Furthermore, even if deleting one or more amino acids from the N-terminus or C-terminus of a polypeptide results in modification or loss of one or more biological functions, other biological activities may still be retained. For example, the ability of a deletion variant to induce and/or to bind antibodies which recognize the secreted form will likely be retained when less than the majority of the residues of the secreted form are removed from the N-terminus or C-terminus. Whether a particular polypeptide lacking N- or C-terminal residues of a protein retains such immunogenic activities can readily be determined by routine methods described herein and otherwise known in the art.

Thus, the invention further includes polypeptide variants which show a biological or functional activity of the polypeptides of the invention (such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune disorders). Such variants include deletions, insertions, inversions, repeats, and substitutions selected according to general rules known in the art so as have little effect on activity.

The present application is directed to nucleic acid molecules at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the nucleic acid sequences disclosed herein, (e.g., encoding a polypeptide having the amino acid sequence of an N and/or C terminal deletion), irrespective of whether they encode a polypeptide having functional activity. This is because even where a particular nucleic acid molecule does not encode a polypeptide having functional activity, one of skill in the art would still know how to use the nucleic acid molecule, for instance, as a hybridization probe or a polymerase chain reaction (PCR) primer. Uses of the nucleic acid molecules of the present invention that do not encode a polypeptide having functional activity include, inter alia, (1) isolating a gene or allelic or splice variants thereof in a cDNA library; (2) in situ hybridization (e.g., “FISH”) to metaphase chromosomal spreads to provide precise chromosomal location of the gene, as described in Verma et al., Human Chromosomes: A Manual of Basic Techniques, Pergamon Press, New York (1988); (3) Northern Blot analysis for detecting mRNA expression in specific tissues (e.g., normal or diseased tissues); and (4) in situ hybridization (e.g., histochemistry) for detecting mRNA expression in specific tissues (e.g., normal or diseased tissues).

Preferred, however, are nucleic acid molecules having sequences at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the nucleic acid sequences disclosed herein, which do, in fact, encode a polypeptide having functional activity. By a polypeptide having “functional activity” is meant, a polypeptide capable of displaying one or more known functional activities associated with a full-length (complete) protein and/or a mature (secreted) protein of the invention. Such functional activities include, but are not limited to, biological activity (such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune diseases and disorders), antigenicity (ability to bind, or compete with a polypeptide of the invention for binding, to an anti-polypeptide of the invention antibody), immunogenicity (ability to generate antibody which binds to a specific polypeptide of the invention), ability to form multimers with polypeptides of the invention, and ability to bind to a receptor or ligand for a polypeptide of the invention.

The functional activity of the polypeptides, and fragments, variants and derivatives of the invention, can be assayed by various methods.

For example, in one embodiment where one is assaying for the ability to bind or compete with a full-length polypeptide of the present invention for binding to an anti-polypetide antibody, various immunoassays known in the art can be used, including but not limited to, competitive and non-competitive assay systems using techniques such as radioimmunoassays, ELISA (enzyme linked immunosorbent assay), “sandwich” immunoassays, immunoradiometric assays, gel diffusion precipitation reactions, immunodiffusion assays, in situ immunoassays (using colloidal gold, enzyme or radioisotope labels, for example), western blots, precipitation reactions, agglutination assays (e.g., gel agglutination assays, hemagglutination assays), complement fixation assays, immunofluorescence assays, protein A assays, and immunoelectrophoresis assays, etc. In one embodiment, antibody binding is detected by detecting a label on the primary antibody. In another embodiment, the primary antibody is detected by detecting binding of a secondary antibody or reagent to the primary antibody. In a further embodiment, the secondary antibody is labeled. Many means are known in the art for detecting binding in an immunoassay and are within the scope of the present invention.

In another embodiment, where a ligand is identified, or the ability of a polypeptide fragment, variant or derivative of the invention to multimerize is being evaluated, binding can be assayed, e.g., by means well-known in the art, such as, for example, reducing and non-reducing gel chromatography, protein affinity chromatography, and affinity blotting. See generally, Phizicky et al., Microbiol. Rev. 59:94-123 (1995). In another embodiment, the ability of physiological correlates of a polypeptide of the present invention to bind to a substrate(s) of the polypeptide of the invention can be routinely assayed using techniques known in the art.

In addition, assays described herein (see Examples) and otherwise known in the art may routinely be applied to measure the ability of polypeptides of the present invention and fragments, variants and derivatives thereof to elicit polypeptide related biological activity (either in vitro or in vivo). Other methods will be known to the skilled artisan and are within the scope of the invention.

Of course, due to the degeneracy of the genetic code, one of ordinary skill in the art will immediately recognize that a large number of the nucleic acid molecules having a sequence at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to, for example, the nucleic acid sequence of the cDNA contained in ATCC™ Deposit No:Z, the nucleic acid sequence referred to in Table 1B (SEQ ID NO:X), the nucleic acid sequence disclosed in Table 1A (e.g., the nucleic acid sequence delineated in columns 7 and 8), the nucleic acid sequence disclosed in Table 2 (e.g., the nucleic acid sequence delineated in columns 8 and 9) or fragments thereof, will encode polypeptides “having functional activity.” In fact, since degenerate variants of any of these nucleotide sequences all encode the same polypeptide, in many instances, this will be clear to the skilled artisan even without performing the above described comparison assay. It will be further recognized in the art that, for such nucleic acid molecules that are not degenerate variants, a reasonable number will also encode a polypeptide having functional activity. This is because the skilled artisan is fully aware of amino acid substitutions that are either less likely or not likely to significantly effect protein function (e.g., replacing one aliphatic amino acid with a second aliphatic amino acid), as further described below.

For example, guidance concerning how to make phenotypically silent amino acid substitutions is provided in Bowie et al., “Deciphering the Message in Protein Sequences: Tolerance to Amino Acid Substitutions,” Science 247:1306-1310 (1990), wherein the authors indicate that there are two main strategies for studying the tolerance of an amino acid sequence to change.

The first strategy exploits the tolerance of amino acid substitutions by natural selection during the process of evolution. By comparing amino acid sequences in different species, conserved amino acids can be identified. These conserved amino acids are likely important for protein function. In contrast, the amino acid positions where substitutions have been tolerated by natural selection indicates that these positions are not critical for protein function. Thus, positions tolerating amino acid substitution could be modified while still maintaining biological activity of the protein.

The second strategy uses genetic engineering to introduce amino acid changes at specific positions of a cloned gene to identify regions critical for protein function. For example, site directed mutagenesis or alanine-scanning mutagenesis (introduction of single alanine mutations at every residue in the molecule) can be used. See Cunningham and Wells, Science 244:1081-1085 (1989). The resulting mutant molecules can then be tested for biological activity.

As the authors state, these two strategies have revealed that proteins are surprisingly tolerant of amino acid substitutions. The authors further indicate which amino acid changes are likely to be permissive at certain amino acid positions in the protein. For example, most buried (within the tertiary structure of the protein) amino acid residues require nonpolar side chains, whereas few features of surface side chains are generally conserved. Moreover, tolerated conservative amino acid substitutions involve replacement of the aliphatic or hydrophobic amino acids Ala, Val, Leu and Ile; replacement of the hydroxyl residues Ser and Thr; replacement of the acidic residues Asp and Glu; replacement of the amide residues Asn and Gln, replacement of the basic residues Lys, Arg, and His; replacement of the aromatic residues Phe, Tyr, and Trp, and replacement of the small-sized amino acids Ala, Ser, Thr, Met, and Gly.

Besides conservative amino acid substitution, variants of the present invention include (i) substitutions with one or more of the non-conserved amino acid residues, where the substituted amino acid residues may or may not be one encoded by the genetic code, or (ii) substitutions with one or more of the amino acid residues having a substituent group, or (iii) fusion of the mature polypeptide with another compound, such as a compound to increase the stability and/or solubility of the polypeptide (for example, polyethylene glycol), (iv) fusion of the polypeptide with additional amino acids, such as, for example, an IgG Fc fusion region peptide, serum albumin (preferably human serum albumin) or a fragment thereof, or leader or secretory sequence, or a sequence facilitating purification, or (v) fusion of the polypeptide with another compound, such as albumin (including but not limited to recombinant albumin (see, e.g., U.S. Pat. No. 5,876,969, issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No. 5,766,883, issued Jun. 16, 1998, herein incorporated by reference in their entirety)). Such variant polypeptides are deemed to be within the scope of those skilled in the art from the teachings herein.

For example, polypeptide variants containing amino acid substitutions of charged amino acids with other charged or neutral amino acids may produce proteins with improved characteristics, such as less aggregation. Aggregation of pharmaceutical formulations both reduces activity and increases clearance due to the aggregate's immunogenic activity. See Pinckard et al., Clin. Exp. Immunol 2.331-340 (1967); Robbins et al., Diabetes 36: 838-845 (1987); Cleland et al., Crit. Rev. Therapeutic Drug Carrier Systems 10:307-377 (1993).

A further embodiment of the invention relates to polypeptides which comprise the amino acid sequence of a polypeptide having an amino acid sequence which contains at least one amino acid substitution, but not more than 50 amino acid substitutions, even more preferably, not more than 40 amino acid substitutions, still more preferably, not more than 30 amino acid substitutions, and still even more preferably, not more than 20 amino acid substitutions from a polypeptide sequence disclosed herein. Of course it is highly preferable for a polypeptide to have an amino acid sequence which, for example, comprises the amino acid sequence of a polypeptide of SEQ ID NO:Y, the amino acid sequence of the mature (e.g., secreted) polypeptide of SEQ ID NO:Y, an amino acid sequence encoded by SEQ ID NO:X, an amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, an amino acid sequence encoded by the complement of SEQ ID NO:X, an amino acid sequence encoded by cDNA contained in ATCC™ Deposit No:Z, and/or the amino acid sequence of a mature (secreted) polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z, or a fragment thereof, which contains, in order of ever-increasing preference, at least one, but not more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 amino acid substitutions.

In specific embodiments, the polypeptides of the invention comprise, or alternatively, consist of, fragments or variants of a reference amino acid sequence selected from: (a) the amino acid sequence of SEQ ID NO:Y or fragments thereof (e.g., the mature form and/or other fragments described herein); (b) the amino acid sequence encoded by SEQ ID NO:X or fragments thereof; (c) the amino acid sequence encoded by the complement of SEQ ID NO:X or fragments thereof; (d) the amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or fragments thereof; and (e) the amino acid sequence encoded by cDNA contained in ATCC™ Deposit No:Z or fragments thereof; wherein the fragments or variants have 1-5, 5-10, 5-25, 5-50, 10-50 or 50-150, amino acid residue additions, substitutions, and/or deletions when compared to the reference amino acid sequence. In preferred embodiments, the amino acid substitutions are conservative. Polynucleotides encoding these polypeptides are also encompassed by the invention.

Polynucleotide and Polypeptide Fragments

The present invention is also directed to polynucleotide fragments of the polynucleotides (nucleic acids) of the invention. In the present invention, a “polynucleotide fragment” refers to a polynucleotide having a nucleic acid sequence which, for example: is a portion of the cDNA contained in ATCC™ Deposit No:Z or the complementary strand thereto; is a portion of the polynucleotide sequence encoding the polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z or the complementary strand thereto; is a portion of the polynucleotide sequence encoding the mature (secreted) polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z or the complementary strand thereto; is a portion of a polynucleotide sequence encoding the mature amino acid sequence as defined in columns 14 and 15 of Table 1A or the complementary strand thereto; is a portion of a polynucleotide sequence encoding the amino acid sequence encoded by the region of SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto; is a portion of the polynucleotide sequence of SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto; is a portion of the polynucleotide sequence in SEQ ID NO:X or the complementary strand thereto; is a polynucleotide sequence encoding a portion of the polypeptide of SEQ ID NO:Y; is a polynucleotide sequence encoding a portion of a polypeptide encoded by SEQ ID NO:X; is a polynucleotide sequence encoding a portion of a polypeptide encoded by the complement of the polynucleotide sequence in SEQ ID NO:X; is a portion of a polynucleotide sequence encoding the amino acid sequence encoded by the region of SEQ ID NO:B as defined in column 6 of Table 1C or the complementary strand thereto; or is a portion of the polynucleotide sequence of SEQ ID NO:B as defined in column 6 of Table 1C or the complementary strand thereto.

The polynucleotide fragments of the invention are preferably at least about 15 nt, and more preferably at least about 20 nt, still more preferably at least about 30 nt, and even more preferably, at least about 40 nt, at least about 50 nt, at least about 75 nt, or at least about 150 nt in length. A fragment “at least 20 nt in length,” for example, is intended to include 20 or more contiguous bases from the cDNA sequence contained in ATCC™ Deposit No:Z, or the nucleotide sequence shown in SEQ ID NO:X or the complementary stand thereto. In this context “about” includes the particularly recited value or a value larger or smaller by several (5, 4, 3, 2, or 1) nucleotides, at either terminus or at both termini. These nucleotide fragments have uses that include, but are not limited to, as diagnostic probes and primers as discussed herein. Of course, larger fragments (e.g., at least 160, 170, 180, 190, 200, 250, 500, 600, 1000, or 2000 nucleotides in length) are also encompassed by the invention.

Moreover, representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a sequence from about nucleotide number 1-50, 51-100, 101-150, 151-200, 201-250, 251-300, 301-350, 351-400, 401-450, 451-500, 501-550, 551-600, 601-650, 651-700, 701-750, 751-800, 801-850, 851-900, 901-950, 951-1000, 1001-1050, 1051-1100, 1101-1150, 1151-1200, 1201-1250, 1251-1300, 1301-1350, 1351-1400, 1401-1450, 1451-1500, 1501-1550, 1551-1600, 1601-1650, 1651-1700, 1701-1750, 1751-1800, 1801-1850, 1851-1900, 1901-1950, 1951-2000, 2001-2050, 2051-2100, 2101-2150, 2151-2200, 2201-2250, 2251-2300, 2301-2350, 2351-2400, 2401-2450, 2451-2500, 2501-2550, 2551-2600, 2601-2650, 2651-2700, 2701-2750, 2751-2800, 2801-2850, 2851-2900, 2901-2950, 2951-3000, 3001-3050, 3051-3100, 3101-3150, 3151-3200, 3201-3250, 3251-3300, 3301-3350, 3351-3400, 3401-3450, 3451-3500, 3501-3550, 3551-3600, 3601-3650, 3651-3700, 3701-3750, 3751-3800, 3801-3850, 3851-3900, 3901-3950, 3951-4000, 4001-4050, 4051-4100, 4101-4150, 4151-4200, 4201-4250, 4251-4300, 4301-4350, 4351-4400, 4401-4450, 4451-4500, 4501-4550, 4551-4600, 4601-4650, 4651-4700, 4701-4750, 4751-4800, 4801-4850, 4851-4900, 4901-4950, 4951-5000, 5001-5050, 5051-5100, 5101-5150, 5151-5200, 5201-5250, 5251-5300, 5301-5350, 5351-5400, 5401-5450, 5451-5500, 5501-5550, 5551-5600, 5601-5650, 5651-5700, 5701-5750, 5751-5800, 5801-5850, 5851-5900, 5901-5950, 5951-6000, 6001-6050, 6051-6100, 6101-6150, 6151-6200, 6201-6250, 6251-6300, 6301-6350, 6351-6400, 6401-6450, 6451-6500, 6501-6550, 6551-6600, 6601-6650, 6651-6700, 6701-6750, 6751-6800, 6801-6850, 6851-6900, 6901-6950, 6951-7000, 7001-7050, 7051-7100, 7101-7150, 7151-7200, 7201-7250, 7251-7300 or 7301 to the end of SEQ ID NO:X, or the complementary strand thereto. In this context “about” includes the particularly recited range or a range larger or smaller by several (5, 4, 3, 2, or 1) nucleotides, at either terminus or at both termini. Preferably, these fragments encode a polypeptide which has a functional activity (e.g., biological activity; such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune diseases and disorders). More preferably, these polynucleotides can be used as probes or primers as discussed herein. Polynucleotides which hybridize to one or more of these polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions are also encompassed by the invention, as are polypeptides encoded by these polynucleotides.

Further representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a sequence from about nucleotide number 1-50, 51-100, 101-150, 151-200, 201-250, 251-300, 301-350, 351-400, 401-450, 451-500, 501-550, 551-600, 601-650, 651-700, 701-750, 751-800, 801-850, 851-900, 901-950, 951-1000, 1001-1050, 1051-1100, 1101-1150, 1151-1200, 1201-1250, 1251-1300, 1301-1350, 1351-1400, 1401-1450, 1451-1500, 1501-1550, 1551-1600, 1601-1650, 1651-1700, 1701-1750, 1751-1800, 1801-1850, 1851-1900, 1901-1950, 1951-2000, 2001-2050, 2051-2100, 2101-2150, 2151-2200, 2201-2250, 2251-2300, 2301-2350, 2351-2400, 2401-2450, 2451-2500, 2501-2550, 2551-2600, 2601-2650, 2651-2700, 2701-2750, 2751-2800, 2801-2850, 2851-2900, 2901-2950, 2951-3000, 3001-3050, 3051-3100, 3101-3150, 3151-3200, 3201-3250, 3251-3300, 3301-3350, 3351-3400, 3401-3450, 3451-3500, 3501-3550, 3551-3600, 3601-3650, 3651-3700, 3701-3750, 3751-3800, 3801-3850, 3851-3900, 3901-3950, 3951-4000, 4001-4050, 4051-4100, 4101-4150, 4151-4200, 4201-4250, 4251-4300, 4301-4350, 4351-4400, 4401-4450, 4451-4500, 4501-4550, 4551-4600, 4601-4650, 4651-4700, 4701-4750, 4751-4800, 4801-4850, 4851-4900, 4901-4950, 4951-5000, 5001-5050, 5051-5100, 5101-5150, 5151-5200, 5201-5250, 5251-5300, 5301-5350, 5351-5400, 5401-5450, 5451-5500, 5501-5550, 5551-5600, 5601-5650, 5651-5700, 5701-5750, 5751-5800, 5801-5850, 5851-5900, 5901-5950, 5951-6000, 6001-6050, 6051-6100, 6101-6150, 6151-6200, 6201-6250, 6251-6300, 6301-6350, 6351-6400, 6401-6450, 6451-6500, 6501-6550, 6551-6600, 6601-6650, 6651-6700, 6701-6750, 6751-6800, 6801-6850, 6851-6900, 6901-6950, 6951-7000, 7001-7050, 7051-7100, 7101-7150, 7151-7200, 7201-7250, 7251-7300 or 7301 to the end of the cDNA sequence contained in ATCC™ Deposit No:Z, or the complementary strand thereto. In this context “about” includes the particularly recited range or a range larger or smaller by several (5, 4, 3, 2, or 1) nucleotides, at either terminus or at both termini. Preferably, these fragments encode a polypeptide which has a functional activity (e.g., biological activity). More preferably, these polynucleotides can be used as probes or primers as discussed herein. Polynucleotides which hybridize to one or more of these polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions are also encompassed by the invention, as are polypeptides encoded by these polynucleotides.

Moreover, representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a nucleic acid sequence comprising one, two, three, four, five, six, seven, eight, nine, ten, or more of the above described polynucleotide fragments of the invention in combination with a polynucleotide sequence delineated in Table 1C column 6. Additional, representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a nucleic acid sequence comprising one, two, three, four, five, six, seven, eight, nine, ten, or more of the above described polynucleotide fragments of the invention in combination with a polynucleotide sequence that is the complementary strand of a sequence delineated in column 6 of Table 1C. In further embodiments, the above-described polynucleotide fragments of the invention comprise, or alternatively consist of, sequences delineated in Table 1C, column 6, and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1C, column 5). In additional embodiments, the above-described polynucleotide fragments of the invention comprise, or alternatively consist of, sequences delineated in Table 1C, column 6, and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated Table 1C, column 6, and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1C, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more fragments of the sequences delineated in column 6 of Table 1C, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1C, column 2) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more fragments of the sequences delineated in column 6 of Table 1C which correspond to the same ATCC™ Deposit No:Z (see Table 1C, column 1), and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A, 1B, or 1C) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more fragments of the sequences delineated in the same row of column 6 of Table 1C, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A, 1B, or 1C) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of the sequence of SEQ ID NO:X are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X (e.g., as described herein) are directly contiguous Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X and the 5′ 10 polynucleotides of the sequence of one of the sequences delineated in column 6 of Table 1C are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C and the 5′ 10 polynucleotides of another sequence in column 6 are directly contiguous. In preferred embodiments, the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1C is directly contiguous with the 5′ 10 polynucleotides of the next sequential exon delineated in Table 1C, column 6. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

In the present invention, a “polypeptide fragment” refers to an amino acid sequence which is a portion of the amino acid sequence contained in SEQ ID NO:Y, is a portion of the mature form of SEQ ID NO:Y as defined in columns 14 and 15 of Table 1A, a portion of an amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, is a portion of an amino acid sequence encoded by the polynucleotide sequence of SEQ ID NO:X, is a portion of an amino acid sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X, is a portion of the amino acid sequence of a mature (secreted) polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z, and/or is a portion of an amino acid sequence encoded by the cDNA contained in ATCC™ Deposit No:Z. Protein (polypeptide) fragments may be “free-standing,” or comprised within a larger polypeptide of which the fragment forms a part or region, most preferably as a single continuous region. Representative examples of polypeptide fragments of the invention, include, for example, fragments comprising, or alternatively consisting of, from about amino acid number 1-20, 21-40, 41-60, 61-80, 81-100, 101-120, 121-140, 141-160, 161-180, 181-200, 201-220, 221-240, 241-260, 261-280, 281-300, 301-320, 321-340, 341-360, 361-380, 381-400, 401-420, 421-440, 441-460, 461-480, 481-500, 501-520, 521-540, 541-560, 561-580, 581-600, 601-620, 621-640, 641-660, 661-680, 681-700, 701-720, 721-740, 741-760, 761-780, 781-800, 801-820, 821-840, 841-860, 861-880, 881-900, 901-920, 921-940, 941-960, 961-980, 981-1000, 1001-1020, 1021-1040, 1041-1060, 1061-1080, 1081-1100, 1101-1120, 1121-1140, 1141-1160, 1161-1180, 1181-1200, 1201-1220, 1221-1240, 1241-1260, 1261-1280, 1281-1300, 1301-1320, 1321-1340, 1341-1360, 1361-1380, 1381-1400, 1401-1420, 1421-1440, or 1441 to the end of the coding region of cDNA and SEQ ID NO: Y. In a preferred embodiment, polypeptide fragments of the invention include, for example, fragments comprising, or alternatively consisting of, from about amino acid number 1-20, 21-40, 41-60, 61-80, 81-100, 101-120, 121-140, 141-160, 161-180, 181-200, 201-220, 221-240, 241-260, 261-280, 281-300, 301-320, 321-340, 341-360, 361-380, 381-400, 401-420, 421-440, 441-460, 461-480, 481-500, 501-520, 521-540, 541-560, 561-580, 581-600, 601-620, 621-640, 641-660, 661-680, 681-700, 701-720, 721-740, 741-760, 761-780, 781-800, 801-820, 821-840, 841-860, 861-880, 881-900, 901-920, 921-940, 941-960, 961-980, 981-1000, 1001-1020, 1021-1040, 1041-1060, 1061-1080, 1081-1100, 1101-1120, 1121-1140, 1141-1160, 1161-1180, 1181-1200, 1201-1220, 1221-1240, 1241-1260, 1261-1280, 1281-1300, 1301-1320, 1321-1340, 1341-1360, 1361-1380, 1381-1400, 1401-1420, 1421-1440, or 1441 to the end of the coding region of SEQ ID NO:Y. Moreover, polypeptide fragments of the invention may be at least about 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 100, 110, 120, 130, 140, or 150 amino acids in length. In this context “about” includes the particularly recited ranges or values, or ranges or values larger or smaller by several (5, 4, 3, 2, or 1) amino acids, at either extreme or at both extremes. Polynucleotides encoding these polypeptide fragments are also encompassed by the invention.

Even if deletion of one or more amino acids from the N-terminus of a protein results in modification of loss of one or more biological functions of the protein, other functional activities (e.g., biological activities; such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune diseases and disorders; ability to multimerize; ability to bind a ligand; antigenic ability useful for production of polypeptide specific antibodies) may still be retained. For example, the ability of shortened muteins to induce and/or bind to antibodies which recognize the complete or mature forms of the polypeptides generally will be retained when less than the majority of the residues of the complete or mature polypeptide are removed from the N-terminus. Whether a particular polypeptide lacking N-terminal residues of a complete polypeptide retains such immunologic activities can readily be determined by routine methods described herein and otherwise known in the art. It is not unlikely that a mutein with a large number of deleted N-terminal amino acid residues may retain some biological or immunogenic activities. In fact, peptides composed of as few as six amino acid residues may often evoke an immune response.

Accordingly, polypeptide fragments include the secreted protein as well as the mature form. Further preferred polypeptide fragments include the secreted protein or the mature form having a continuous series of deleted residues from the amino or the carboxy terminus, or both. For example, any number of amino acids, ranging from 1-60, can be deleted from the amino terminus of either the secreted polypeptide or the mature form. Similarly, any number of amino acids, ranging from 1-30, can be deleted from the carboxy terminus of the secreted protein or mature form. Furthermore, any combination of the above amino and carboxy terminus deletions are preferred. Similarly, polynucleotides encoding these polypeptide fragments are also preferred.

The present invention further provides polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of a polypeptide disclosed herein (e.g., a polypeptide of SEQ ID NO:Y, a polypeptide as defined in columns 14 and 15 of Table 1A, a polypeptide encoded by the polynucleotide sequence contained in SEQ ID NO:X or the complement thereof, a polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, a polypeptide encoded by the portion of SEQ ID NO:B as defined in column 6 of Table 1C, a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z, and/or a mature polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z). In particular, N-terminal deletions may be described by the general formula III-q, where q is a whole integer representing the total number of amino acid residues in a polypeptide of the invention (e.g., the polypeptide disclosed in SEQ ID NO:Y, the mature (secreted) portion of SEQ ID NO:Y as defined in columns 14 and 15 of Table 1A, or the polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2), and m is defined as any integer ranging from 2 to q-6. Polynucleotides encoding these polypeptides are also encompassed by the invention.

The present invention further provides polypeptides having one or more residues from the carboxy terminus of the amino acid sequence of a polypeptide disclosed herein (e.g., a polypeptide of SEQ ID NO:Y, the mature (secreted) portion of SEQ ID NO:Y as defined in columns 14 and 15 of Table 1A, a polypeptide encoded by the polynucleotide sequence contained in SEQ ID NO:X, a polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, a polypeptide encoded by the portion of SEQ ID NO:B as defined in column 6 of Table 1C, a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z, and/or a mature polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z). In particular, C-terminal deletions may be described by the general formula I-n, where n is any whole integer ranging from 6 to q-1, and where n corresponds to the position of amino acid residue in a polypeptide of the invention. Polynucleotides encoding these polypeptides are also encompassed by the invention.

In addition, any of the above described N- or C-terminal deletions can be combined to produce a N- and C-terminal deleted polypeptide. The invention also provides polypeptides having one or more amino acids deleted from both the amino and the carboxyl termini, which may be described generally as having residues m-n of a polypeptide encoded by SEQ ID NO:X (e.g., including, but not limited to, the preferred polypeptide disclosed as SEQ ID NO:Y, the mature (secreted) portion of SEQ ID NO:Y as defined in columns 14 and 15 of Table 1A, and the polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2), the cDNA contained in ATCC™ Deposit No:Z, and/or the complement thereof, where n and m are integers as described above. Polynucleotides encoding these polypeptides are also encompassed by the invention.

Also as mentioned above, even if deletion of one or more amino acids from the C-terminus of a protein results in modification of loss of one or more biological functions of the protein, other functional activities (e.g., biological activities such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune diseases and disorders; ability to multimerize; ability to bind a ligand; antigenic ability useful for production of polypeptide specific antibodies) may still be retained. For example the ability of the shortened mutein to induce and/or bind to antibodies which recognize the complete or mature forms of the polypeptide generally will be retained when less than the majority of the residues of the complete or mature polypeptide are removed from the C-terminus. Whether a particular polypeptide lacking C-terminal residues of a complete polypeptide retains such immunologic activities can readily be determined by routine methods described herein and otherwise known in the art. It is not unlikely that a mutein with a large number of deleted C-terminal amino acid residues may retain some biological or immunogenic activities. In fact, peptides composed of as few as six amino acid residues may often evoke an immune response.

The present application is also directed to proteins containing polypeptides at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to a polypeptide sequence set forth herein. In preferred embodiments, the application is directed to proteins containing polypeptides at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to polypeptides having the amino acid sequence of the specific N- and C-terminal deletions. Polynucleotides encoding these polypeptides are also encompassed by the invention.

Any polypeptide sequence encoded by, for example, the polynucleotide sequences set forth as SEQ ID NO:X or the complement thereof, (presented, for example, in Tables 1A and 2), the cDNA contained in ATCC™ Deposit No:Z, or the polynucleotide sequence as defined in column 6 of Table 1C, may be analyzed to determine certain preferred regions of the polypeptide. For example, the amino acid sequence of a polypeptide encoded by a polynucleotide sequence of SEQ ID NO:X (e.g., the polypeptide of SEQ ID NO:Y and the polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2) or the cDNA contained in ATCC™ Deposit No:Z may be analyzed using the default parameters of the DNASTAR computer algorithm (DNASTAR, Inc., 1228 S. Park St., Madison, Wis. 53715 USA).

Polypeptide regions that may be routinely obtained using the DNASTAR computer algorithm include, but are not limited to, Garnier-Robson alpha-regions, beta-regions, turn-regions, and coil-regions; Chou-Fasman alpha-regions, beta-regions, and turn-regions; Kyte-Doolittle hydrophilic regions and hydrophobic regions; Eisenberg alpha- and beta-amphipathic regions; Karplus-Schulz flexible regions; Emini surface-forming regions; and Jameson-Wolf regions of high antigenic index. Among highly preferred polynucleotides of the invention in this regard are those that encode polypeptides comprising regions that combine several structural features, such as several (e.g., 1, 2, 3 or 4) of the features set out above.

Additionally, Kyte-Doolittle hydrophilic regions and hydrophobic regions, Emini surface-forming regions, and Jameson-Wolf regions of high antigenic index (i.e., containing four or more contiguous amino acids having an antigenic index of greater than or equal to 1.5, as identified using the default parameters of the Jameson-Wolf program) can routinely be used to determine polypeptide regions that exhibit a high degree of potential for antigenicity. Regions of high antigenicity are determined from data by DNASTAR analysis by choosing values which represent regions of the polypeptide which are likely to be exposed on the surface of the polypeptide in an environment in which antigen recognition may occur in the process of initiation of an immune response.

Preferred polypeptide fragments of the invention are fragments comprising, or alternatively, consisting of, an amino acid sequence that displays a functional activity (e.g. biological activity such as, for example, activity useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating immune diseases and disorders; ability to multimerize; ability to bind a ligand; antigenic ability useful for production of polypeptide specific antibodies) of the polypeptide sequence of which the amino acid sequence is a fragment. By a polypeptide displaying a “functional activity” is meant a polypeptide capable of one or more known functional activities associated with a full-length protein, such as, for example, biological activity, antigenicity, immunogenicity, and/or multimerization, as described herein.

Other preferred polypeptide fragments are biologically active fragments. Biologically active fragments are those exhibiting activity similar, but not necessarily identical, to an activity of the polypeptide of the present invention. The biological activity of the fragments may include an improved desired activity, or a decreased undesirable activity.

In preferred embodiments, polypeptides of the invention comprise, or alternatively consist of, one, two, three, four, five or more of the antigenic fragments of the polypeptide of SEQ ID NO:Y, or portions thereof. Polynucleotides encoding these polypeptides are also encompassed by the invention.

Epitopes and Antibodies

The present invention encompasses polypeptides comprising, or alternatively consisting of, an epitope of: the polypeptide sequence shown in SEQ ID NO:Y; a polypeptide sequence encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2; the polypeptide sequence encoded by the portion of SEQ ID NO:B as defined in column 6 of Table 1C or the complement thereto; the polypeptide sequence encoded by the cDNA contained in ATCC™ Deposit No:Z; or the polypeptide sequence encoded by a polynucleotide that hybridizes to the sequence of SEQ ID NO:X, the complement of the sequence of SEQ ID NO:X, the complement of a portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, or the cDNA sequence contained in ATCC™ Deposit No:Z under stringent hybridization conditions or alternatively, under lower stringency hybridization as defined supra. The present invention further encompasses polynucleotide sequences encoding an epitope of a polypeptide sequence of the invention (such as, for example, the sequence disclosed in SEQ ID NO:X, or a fragment thereof), polynucleotide sequences of the complementary strand of a polynucleotide sequence encoding an epitope of the invention, and polynucleotide sequences which hybridize to the complementary strand under stringent hybridization conditions or alternatively, under lower stringency hybridization conditions defined supra.

The term “epitopes,” as used herein, refers to portions of a polypeptide having antigenic or immunogenic activity in an animal, preferably a mammal, and most preferably in a human. In a preferred embodiment, the present invention encompasses a polypeptide comprising an epitope, as well as the polynucleotide encoding this polypeptide. An “immunogenic epitope,” as used herein, is defined as a portion of a protein that elicits an antibody response in an animal, as determined by any method known in the art, for example, by the methods for generating antibodies described infra. (See, for example, Geysen et al., Proc. Natl. Acad. Sci. USA 81:3998-4002 (1983)). The term “antigenic epitope,” as used herein, is defined as a portion of a protein to which an antibody can immunospecifically bind its antigen as determined by any method well known in the art, for example, by the immunoassays described herein Immunospecific binding excludes non-specific binding but does not necessarily exclude cross-reactivity with other antigens. Antigenic epitopes need not necessarily be immunogenic.

Fragments which function as epitopes may be produced by any conventional means. (See, e.g., Houghten, R. A., Proc. Natl. Acad. Sci. USA 82:5131-5135 (1985) further described in U.S. Pat. No. 4,631,211.)

In the present invention, antigenic epitopes preferably contain a sequence of at least 4, at least 5, at least 6, at least 7, more preferably at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 20, at least 25, at least 30, at least 40, at least 50, and, most preferably, between about 15 to about 30 amino acids. Preferred polypeptides comprising immunogenic or antigenic epitopes are at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 amino acid residues in length. Additional non-exclusive preferred antigenic epitopes include the antigenic epitopes disclosed herein, as well as portions thereof. Antigenic epitopes are useful, for example, to raise antibodies, including monoclonal antibodies, that specifically bind the epitope. Preferred antigenic epitopes include the antigenic epitopes disclosed herein, as well as any combination of two, three, four, five or more of these antigenic epitopes. Antigenic epitopes can be used as the target molecules in immunoassays. (See, for instance, Wilson et al., Cell 37:767-778 (1984); Sutcliffe et al., Science 219:660-666 (1983)).

Non-limiting examples of epitopes of polypeptides that can be used to generate antibodies of the invention include a polypeptide comprising, or alternatively consisting of, at least one, two, three, four, five, six or more of the portion(s) of SEQ ID NO:Y specified in Table 1B. These polypeptide fragments have been determined to bear antigenic epitopes of the proteins of the invention by the analysis of the Jameson-Wolf antigenic index that is included in the DNAStar suite of computer programs. By “comprise” it is intended that a polypeptide contains at least one, two, three, four, five, six or more of the portion(s) of SEQ ID NO:Y shown in Table 1B, but it may contain additional flanking residues on either the amino or carboxyl termini of the recited portion. Such additional flanking sequences are preferably sequences naturally found adjacent to the portion; i.e., contiguous sequence shown in SEQ ID NO:Y. The flanking sequence may, however, be sequences from a heterologous polypeptide, such as from another protein described herein or from a heterologous polypeptide not described herein. In particular embodiments, epitope portions of a polypeptide of the invention comprise one, two, three, or more of the portions of SEQ ID NO:Y shown in Table 1B.

Similarly, immunogenic epitopes can be used, for example, to induce antibodies according to methods well known in the art. See, for instance, Sutcliffe et al., supra; Wilson et al., supra; Chow et al., Proc. Natl. Acad. Sci. USA 82:910-914; and Bittle et al., J. Gen. Virol. 66:2347-2354 (1985). Preferred immunogenic epitopes include the immunogenic epitopes disclosed herein, as well as any combination of two, three, four, five or more of these immunogenic epitopes. The polypeptides comprising one or more immunogenic epitopes may be presented for eliciting an antibody response together with a carrier protein, such as an albumin, to an animal system (such as rabbit or mouse), or, if the polypeptide is of sufficient length (at least about 25 amino acids), the polypeptide may be presented without a carrier. However, immunogenic epitopes comprising as few as 8 to 10 amino acids have been shown to be sufficient to raise antibodies capable of binding to, at the very least, linear epitopes in a denatured polypeptide (e.g., in Western blotting).

Epitope-bearing polypeptides of the present invention may be used to induce antibodies according to methods well known in the art including, but not limited to, in vivo immunization, in vitro immunization, and phage display methods. See, e.g., Sutcliffe et al., supra; Wilson et al., supra, and Bittle et al., J. Gen. Virol., 66:2347-2354 (1985). If in vivo immunization is used, animals may be immunized with free peptide; however, anti-peptide antibody titer may be boosted by coupling the peptide to a macromolecular carrier, such as keyhole limpet hemacyanin (KLH) or tetanus toxoid. For instance, peptides containing cysteine residues may be coupled to a carrier using a linker such as maleimidobenzoyl-N-hydroxysuccinimide ester (MBS), while other peptides may be coupled to carriers using a more general linking agent such as glutaraldehyde. Animals such as rabbits, rats and mice are immunized with either free or carrier-coupled peptides, for instance, by intraperitoneal and/or intradermal injection of emulsions containing about 100 μg of peptide or carrier protein and Freund's adjuvant or any other adjuvant known for stimulating an immune response. Several booster injections may be needed, for instance, at intervals of about two weeks, to provide a useful titer of anti-peptide antibody which can be detected, for example, by ELISA assay using free peptide adsorbed to a solid surface. The titer of anti-peptide antibodies in serum from an immunized animal may be increased by selection of anti-peptide antibodies, for instance, by adsorption to the peptide on a solid support and elution of the selected antibodies according to methods well known in the art.

As one of skill in the art will appreciate, and as discussed above, the polypeptides of the present invention (e.g., those comprising an immunogenic or antigenic epitope) can be fused to heterologous polypeptide sequences. For example, polypeptides of the present invention (including fragments or variants thereof), may be fused with the constant domain of immunoglobulins (IgA, IgE, IgG, IgM), or portions thereof (CH1, CH2, CH3, or any combination thereof and portions thereof, resulting in chimeric polypeptides. By way of another non-limiting example, polypeptides and/or antibodies of the present invention (including fragments or variants thereof) may be fused with albumin (including but not limited to recombinant human serum albumin or fragments or variants thereof (see, e.g., U.S. Pat. No. 5,876,969, issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No. 5,766,883, issued Jun. 16, 1998, herein incorporated by reference in their entirety)). In a preferred embodiment, polypeptides and/or antibodies of the present invention (including fragments or variants thereof) are fused with the mature form of human serum albumin (i.e., amino acids 1-585 of human serum albumin as shown in FIGS. 1 and 2 of EP Patent 0 322 094) which is herein incorporated by reference in its entirety. In another preferred embodiment, polypeptides and/or antibodies of the present invention (including fragments or variants thereof) are fused with polypeptide fragments comprising, or alternatively consisting of, amino acid residues 1-z of human serum albumin, where z is an integer from 369 to 419, as described in U.S. Pat. No. 5,766,883 herein incorporated by reference in its entirety. Polypeptides and/or antibodies of the present invention (including fragments or variants thereof) may be fused to either the N- or C-terminal end of the heterologous protein (e.g., immunoglobulin Fc polypeptide or human serum albumin polypeptide). Polynucleotides encoding fusion proteins of the invention are also encompassed by the invention.

Such fusion proteins as those described above may facilitate purification and may increase half-life in vivo. This has been shown for chimeric proteins consisting of the first two domains of the human CD4-polypeptide and various domains of the constant regions of the heavy or light chains of mammalian immunoglobulins. See, e.g., EP 394,827; Traunecker et al., Nature, 331:84-86 (1988). Enhanced delivery of an antigen across the epithelial barrier to the immune system has been demonstrated for antigens (e.g., insulin) conjugated to an FcRn binding partner such as IgG or Fc fragments (see, e.g., PCT Publications WO 96/22024 and WO 99/04813). IgG fusion proteins that have a disulfide-linked dimeric structure due to the IgG portion disulfide bonds have also been found to be more efficient in binding and neutralizing other molecules than monomeric polypeptides or fragments thereof alone. See, e.g., Fountoulakis et al., J. Biochem., 270:3958-3964 (1995). Nucleic acids encoding the above epitopes can also be recombined with a gene of interest as an epitope tag (e.g., the hemagglutinin (HA) tag or flag tag) to aid in detection and purification of the expressed polypeptide. For example, a system described by Janknecht et al. allows for the ready purification of non-denatured fusion proteins expressed in human cell lines (Janknecht et al., 1991, Proc. Natl. Acad. Sci. USA 88:8972-897). In this system, the gene of interest is subcloned into a vaccinia recombination plasmid such that the open reading frame of the gene is translationally fused to an amino-terminal tag consisting of six histidine residues. The tag serves as a matrix binding domain for the fusion protein. Extracts from cells infected with the recombinant vaccinia virus are loaded onto Ni2+ nitriloacetic acid-agarose column and histidine-tagged proteins can be selectively eluted with imidazole-containing buffers.

Fusion Proteins

Any polypeptide of the present invention can be used to generate fusion proteins. For example, the polypeptide of the present invention, when fused to a second protein, can be used as an antigenic tag. Antibodies raised against the polypeptide of the present invention can be used to indirectly detect the second protein by binding to the polypeptide. Moreover, because secreted proteins target cellular locations based on trafficking signals, polypeptides of the present invention which are shown to be secreted can be used as targeting molecules once fused to other proteins.

Examples of domains that can be fused to polypeptides of the present invention include not only heterologous signal sequences, but also other heterologous functional regions. The fusion does not necessarily need to be direct, but may occur through linker sequences.

In certain preferred embodiments, proteins of the invention are fusion proteins comprising an amino acid sequence that is an N and/or C-terminal deletion of a polypeptide of the invention. In preferred embodiments, the invention is directed to a fusion protein comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to a polypeptide sequence of the invention. Polynucleotides encoding these proteins are also encompassed by the invention.

Moreover, fusion proteins may also be engineered to improve characteristics of the polypeptide of the present invention. For instance, a region of additional amino acids, particularly charged amino acids, may be added to the N-terminus of the polypeptide to improve stability and persistence during purification from the host cell or subsequent handling and storage. Also, peptide moieties may be added to the polypeptide to facilitate purification. Such regions may be removed prior to final preparation of the polypeptide. The addition of peptide moieties to facilitate handling of polypeptides are familiar and routine techniques in the art.

As one of skill in the art will appreciate that, as discussed above, polypeptides of the present invention, and epitope-bearing fragments thereof, can be combined with heterologous polypeptide sequences. For example, the polypeptides of the present invention may be fused with heterologous polypeptide sequences, for example, the polypeptides of the present invention may be fused with the constant domain of immunoglobulins (IgA, IgE, IgG, IgM) or portions thereof (CH1, CH2, CH3, and any combination thereof, including both entire domains and portions thereof), or albumin (including, but not limited to, native or recombinant human albumin or fragments or variants thereof (see, e.g., U.S. Pat. No. 5,876,969, issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No. 5,766,883, issued Jun. 16, 1998, herein incorporated by reference in their entirety)), resulting in chimeric polypeptides. For example, EP-A-0 464 533 (Canadian counterpart 2045869) discloses fusion proteins comprising various portions of constant region of immunoglobulin molecules together with another human protein or part thereof. In many cases, the Fc part in a fusion protein is beneficial in therapy and diagnosis, and thus can result in, for example, improved pharmacokinetic properties (EP-A 0232 262). Alternatively, deleting the Fc part after the fusion protein has been expressed, detected, and purified, would be desired. For example, the Fc portion may hinder therapy and diagnosis if the fusion protein is used as an antigen for immunizations. In drug discovery, for example, human proteins, such as hIL-5, have been fused with Fc portions for the purpose of high-throughput screening assays to identify antagonists of hIL-5. See, D. Bennett et al., J. Molecular Recognition 8:52-58 (1995); K. Johanson et al., J. Biol. Chem. 270:9459-9471 (1995).

Moreover, the polypeptides of the present invention can be fused to marker sequences, such as a polypeptide which facilitates purification of the fused polypeptide. In preferred embodiments, the marker amino acid sequence is a hexa-histidine peptide, such as the tag provided in a pQE vector (QIAGEN, Inc., 9259 Eton Avenue, Chatsworth, Calif., 91311), among others, many of which are commercially available. As described in Gentz et al., Proc. Natl. Acad. Sci. USA 86:821-824 (1989), for instance, hexa-histidine provides for convenient purification of the fusion protein. Another peptide tag useful for purification, the “HA” tag, corresponds to an epitope derived from the influenza hemagglutinin protein (Wilson et al., Cell 37:767 (1984)).

Additional fusion proteins of the invention may be generated through the techniques of gene-shuffling, motif-shuffling, exon-shuffling, and/or codon-shuffling (collectively referred to as “DNA shuffling”). DNA shuffling may be employed to modulate the activities of polypeptides of the invention, such methods can be used to generate polypeptides with altered activity, as well as agonists and antagonists of the polypeptides. See, generally, U.S. Pat. Nos. 5,605,793; 5,811,238; 5,830,721; 5,834,252; and 5,837,458, and Patten et al., Curr. Opinion Biotechnol. 8:724-33 (1997); Harayama, Trends Biotechnol. 16(2):76-82 (1998); Hansson, et al., J. Mol. Biol. 287:265-76 (1999); and Lorenzo and Blasco, Biotechniques 24(2):308-13 (1998) (each of these patents and publications are hereby incorporated by reference in its entirety). In one embodiment, alteration of polynucleotides corresponding to SEQ ID NO:X and the polypeptides encoded by these polynucleotides may be achieved by DNA shuffling. DNA shuffling involves the assembly of two or more DNA segments by homologous or site-specific recombination to generate variation in the polynucleotide sequence. In another embodiment, polynucleotides of the invention, or the encoded polypeptides, may be altered by being subjected to random mutagenesis by error-prone PCR, random nucleotide insertion or other methods prior to recombination. In another embodiment, one or more components, motifs, sections, parts, domains, fragments, etc., of a polynucleotide encoding a polypeptide of the invention may be recombined with one or more components, motifs, sections, parts, domains, fragments, etc. of one or more heterologous molecules.

Thus, any of these above fusions can be engineered using the polynucleotides or the polypeptides of the present invention.

Recombinant and Synthetic Production of Polypeptides of the Invention

The present invention also relates to vectors containing the polynucleotide of the present invention, host cells, and the production of polypeptides by synthetic and recombinant techniques. The vector may be, for example, a phage, plasmid, viral, or retroviral vector. Retroviral vectors may be replication competent or replication defective. In the latter case, viral propagation generally will occur only in complementing host cells.

The polynucleotides of the invention may be joined to a vector containing a selectable marker for propagation in a host. Generally, a plasmid vector is introduced in a precipitate, such as a calcium phosphate precipitate, or in a complex with a charged lipid. If the vector is a virus, it may be packaged in vitro using an appropriate packaging cell line and then transduced into host cells.

The polynucleotide insert should be operatively linked to an appropriate promoter, such as the phage lambda PL promoter, the E. coli lac, trp, phoA and tac promoters, the SV40 early and late promoters and promoters of retroviral LTRs, to name a few. Other suitable promoters will be known to the skilled artisan. The expression constructs will further contain sites for transcription initiation, termination, and, in the transcribed region, a ribosome binding site for translation. The coding portion of the transcripts expressed by the constructs will preferably include a translation initiating codon at the beginning and a termination codon (UAA, UGA or UAG) appropriately positioned at the end of the polypeptide to be translated.

As indicated, the expression vectors will preferably include at least one selectable marker. Such markers include dihydrofolate reductase, G418, glutamine synthase, or neomycin resistance for eukaryotic cell culture, and tetracycline, kanamycin or ampicillin resistance genes for culturing in E. coli and other bacteria. Representative examples of appropriate hosts include, but are not limited to, bacterial cells, such as E. coli, Streptomyces and Salmonella typhimurium cells; fungal cells, such as yeast cells (e.g., Saccharomyces cerevisiae or Pichia pastoris (ATCC™ Accession No. 201178)); insect cells such as Drosophila S2 and Spodoptera 519 cells; animal cells such as CHO, COS, 293, and Bowes melanoma cells; and plant cells. Appropriate culture mediums and conditions for the above-described host cells are known in the art.

Among vectors preferred for use in bacteria include pQE70, pQE60 and pQE-9, available from QIAGEN, Inc.; pBLUESCRIPT™ vectors, Phagescript vectors, pNH8A, pNH16a, pNH18A, pNH46A, available from STRATAGENE™ mCloning Systems, Inc.; and ptrc99a, pKK223-3, pKK233-3, pDR540, pRIT5 available from PHARMACIA™ Biotech, Inc. Among preferred eukaryotic vectors are pWLNEO, pSV2CAT, pOG44, pXT1 and pSG available from STRATAGENE™; and pSVK3, pBPV, pMSG and pSVL available from PHARMACIAT™. Preferred expression vectors for use in yeast systems include, but are not limited to pYES2, pYD1, pTEF1/Zeo, pYES2/GS, pPICZ, pGAPZ, pGAPZalph, pPIC9, pPIC3.5, pHIL-D2, pHIL-S1, pPIC3.5K, pPIC9K, and PAO815 (all available from Invitrogen, Carlbad, Calif.). Other suitable vectors will be readily apparent to the skilled artisan.

Vectors which use glutamine synthase (GS) or DHFR as the selectable markers can be amplified in the presence of the drugs methionine sulphoximine or methotrexate, respectively. An advantage of glutamine synthase based vectors are the availability of cell lines (e.g., the murine myeloma cell line, NS0) which are glutamine synthase negative. Glutamine synthase expression systems can also function in glutamine synthase expressing cells (e.g., Chinese Hamster Ovary (CHO) cells) by providing additional inhibitor to prevent the functioning of the endogenous gene. A glutamine synthase expression system and components thereof are detailed in PCT publications: WO87/04462; WO86/05807; WO89/01036; WO89/10404; and WO91/06657, which are hereby incorporated in their entireties by reference herein. Additionally, glutamine synthase expression vectors can be obtained from Lonza Biologics, Inc. (Portsmouth, N.H.). Expression and production of monoclonal antibodies using a GS expression system in murine myeloma cells is described in Bebbington et al., Bio/technology 10:169 (1992) and in Biblia and Robinson Biotechnol. Prog. 11:1 (1995) which are herein incorporated by reference.

The present invention also relates to host cells containing the above-described vector constructs described herein, and additionally encompasses host cells containing nucleotide sequences of the invention that are operably associated with one or more heterologous control regions (e.g., promoter and/or enhancer) using techniques known of in the art. The host cell can be a higher eukaryotic cell, such as a mammalian cell (e.g., a human derived cell), or a lower eukaryotic cell, such as a yeast cell, or the host cell can be a prokaryotic cell, such as a bacterial cell. A host strain may be chosen which modulates the expression of the inserted gene sequences, or modifies and processes the gene product in the specific fashion desired. Expression from certain promoters can be elevated in the presence of certain inducers; thus expression of the genetically engineered polypeptide may be controlled. Furthermore, different host cells have characteristics and specific mechanisms for the translational and post-translational processing and modification (e.g., phosphorylation, cleavage) of proteins. Appropriate cell lines can be chosen to ensure the desired modifications and processing of the foreign protein expressed.

Introduction of the nucleic acids and nucleic acid constructs of the invention into the host cell can be effected by calcium phosphate transfection, DEAE-dextran mediated transfection, cationic lipid-mediated transfection, electroporation, transduction, infection, or other methods. Such methods are described in many standard laboratory manuals, such as Davis et al., Basic Methods In Molecular Biology (1986). It is specifically contemplated that the polypeptides of the present invention may in fact be expressed by a host cell lacking a recombinant vector.

In addition to encompassing host cells containing the vector constructs discussed herein, the invention also encompasses primary, secondary, and immortalized host cells of vertebrate origin, particularly mammalian origin, that have been engineered to delete or replace endogenous genetic material (e.g., the coding sequence), and/or to include genetic material (e.g., heterologous polynucleotide sequences) that is operably associated with polynucleotides of the invention, and which activates, alters, and/or amplifies endogenous polynucleotides. For example, techniques known in the art may be used to operably associate heterologous control regions (e.g., promoter and/or enhancer) and endogenous polynucleotide sequences via homologous recombination (see, e.g., U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication Number WO 96/29411; International Publication Number WO 94/12650; Koller et al., Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); and Zijlstra et al., Nature 342:435-438 (1989), the disclosures of each of which are incorporated by reference in their entireties).

Polypeptides of the invention can be recovered and purified from recombinant cell cultures by well-known methods including ammonium sulfate or ethanol precipitation, acid extraction, anion or cation exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxylapatite chromatography and lectin chromatography. Most preferably, high performance liquid chromatography (“HPLC”) is employed for purification.

Polypeptides of the present invention can also be recovered from: products purified from natural sources, including bodily fluids, tissues and cells, whether directly isolated or cultured; products of chemical synthetic procedures; and products produced by recombinant techniques from a prokaryotic or eukaryotic host, including, for example, bacterial, yeast, higher plant, insect, and mammalian cells. Depending upon the host employed in a recombinant production procedure, the polypeptides of the present invention may be glycosylated or may be non-glycosylated. In addition, polypeptides of the invention may also include an initial modified methionine residue, in some cases as a result of host-mediated processes. Thus, it is well known in the art that the N-terminal methionine encoded by the translation initiation codon generally is removed with high efficiency from any protein after translation in all eukaryotic cells. While the N-terminal methionine on most proteins also is efficiently removed in most prokaryotes, for some proteins, this prokaryotic removal process is inefficient, depending on the nature of the amino acid to which the N-terminal methionine is covalently linked.

In one embodiment, the yeast Pichia pastoris is used to express polypeptides of the invention in a eukaryotic system. Pichia pastoris is a methylotrophic yeast which can metabolize methanol as its sole carbon source. A main step in the methanol metabolization pathway is the oxidation of methanol to formaldehyde using O₂. This reaction is catalyzed by the enzyme alcohol oxidase. In order to metabolize methanol as its sole carbon source, Pichia pastoris must generate high levels of alcohol oxidase due, in part, to the relatively low affinity of alcohol oxidase for O₂. Consequently, in a growth medium depending on methanol as a main carbon source, the promoter region of one of the two alcohol oxidase genes (AOX1) is highly active. In the presence of methanol, alcohol oxidase produced from the AOX1 gene comprises up to approximately 30% of the total soluble protein in Pichia pastoris. See Ellis, S. B., et al., Mol. Cell. Biol. 5:1111-21 (1985); Koutz, P. J, et al., Yeast 5:167-77 (1989); Tschopp, J. F., et al., Nucl. Acids Res. 15:3859-76 (1987). Thus, a heterologous coding sequence, such as, for example, a polynucleotide of the present invention, under the transcriptional regulation of all or part of the AOX1 regulatory sequence is expressed at exceptionally high levels in Pichia yeast grown in the presence of methanol.

In one example, the plasmid vector pPIC9K is used to express DNA encoding a polypeptide of the invention, as set forth herein, in a Pichea yeast system essentially as described in “Pichia Protocols: Methods in Molecular Biology,” D. R. Higgins and J. Cregg, eds. The Humana Press, Totowa, N.J., 1998. This expression vector allows expression and secretion of a polypeptide of the invention by virtue of the strong AOX1 promoter linked to the Pichia pastoris alkaline phosphatase (PHO) secretory signal peptide (i.e., leader) located upstream of a multiple cloning site.

Many other yeast vectors could be used in place of pPIC9K, such as, pYES2, pYD1, pTEF1/Zeo, pYES2/GS, pPICZ, pGAPZ, pGAPZalpha, pPIC9, pPIC3.5, pHIL-D2, pHIL-S1, pPIC3.5K, and PAO815, as one skilled in the art would readily appreciate, as long as the proposed expression construct provides appropriately located signals for transcription, translation, secretion (if desired), and the like, including an in-frame AUG as required.

In another embodiment, high-level expression of a heterologous coding sequence, such as, for example, a polynucleotide of the present invention, may be achieved by cloning the heterologous polynucleotide of the invention into an expression vector such as, for example, pGAPZ or pGAPZalpha, and growing the yeast culture in the absence of methanol.

In addition to encompassing host cells containing the vector constructs discussed herein, the invention also encompasses primary, secondary, and immortalized host cells of vertebrate origin, particularly mammalian origin, that have been engineered to delete or replace endogenous genetic material (e.g., coding sequence), and/or to include genetic material (e.g., heterologous polynucleotide sequences) that is operably associated with polynucleotides of the invention, and which activates, alters, and/or amplifies endogenous polynucleotides. For example, techniques known in the art may be used to operably associate heterologous control regions (e.g., promoter and/or enhancer) and endogenous polynucleotide sequences via homologous recombination (see, e.g., U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication No. WO 96/29411, published Sep. 26, 1996; International Publication No. WO 94/12650, published Aug. 4, 1994; Koller et al., Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); and Zijlstra et al., Nature 342:435-438 (1989), the disclosures of each of which are incorporated by reference in their entireties).

In addition, polypeptides of the invention can be chemically synthesized using techniques known in the art (e.g., see Creighton, 1983, Proteins: Structures and Molecular Principles, W.H. Freeman & Co., N.Y., and Hunkapiller et al., Nature, 310:105-111 (1984)). For example, a polypeptide corresponding to a fragment of a polypeptide can be synthesized by use of a peptide synthesizer. Furthermore, if desired, nonclassical amino acids or chemical amino acid analogs can be introduced as a substitution or addition into the polypeptide sequence. Non-classical amino acids include, but are not limited to, to the D-isomers of the common amino acids, 2,4-diaminobutyric acid, a-amino isobutyric acid, 4-aminobutyric acid, Abu, 2-amino butyric acid, g-Abu, e-Ahx, 6-amino hexanoic acid, Aib, 2-amino isobutyric acid, 3-amino propionic acid, ornithine, norleucine, norvaline, hydroxyproline, sarcosine, citrulline, homocitrulline, cysteic acid, t-butylglycine, t-butylalanine, phenylglycine, cyclohexylalanine, b-alanine, fluoro-amino acids, designer amino acids such as b-methyl amino acids, Ca-methyl amino acids, Na-methyl amino acids, and amino acid analogs in general. Furthermore, the amino acid can be D (dextrorotary) or L (levorotary).

The invention encompasses polypeptides of the present invention which are differentially modified during or after translation, e.g., by glycosylation, acetylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, linkage to an antibody molecule or other cellular ligand, etc. Any of numerous chemical modifications may be carried out by known techniques, including but not limited, to specific chemical cleavage by cyanogen bromide, trypsin, chymotrypsin, papain, V8 protease, NaBH₄; acetylation, formylation, oxidation, reduction; metabolic synthesis in the presence of tunicamycin; etc.

Additional post-translational modifications encompassed by the invention include, for example, e.g., N-linked or O-linked carbohydrate chains, processing of N-terminal or C-terminal ends), attachment of chemical moieties to the amino acid backbone, chemical modifications of N-linked or O-linked carbohydrate chains, and addition or deletion of an N-terminal methionine residue as a result of procaryotic host cell expression. The polypeptides may also be modified with a detectable label, such as an enzymatic, fluorescent, isotopic or affinity label to allow for detection and isolation of the protein.

Examples of suitable enzymes include horseradish peroxidase, alkaline phosphatase, beta-galactosidase, or acetylcholinesterase; examples of suitable prosthetic group complexes include streptavidin/biotin and avidin/biotin; examples of suitable fluorescent materials include umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; an example of a luminescent material includes luminol; examples of bioluminescent materials include luciferase, luciferin, and aequorin; and examples of suitable radioactive material include iodine (¹²¹I, ¹²³I, ¹²⁵I, ¹³¹I), carbon (¹⁴C), sulfur (³⁵S), tritium (³H), indium (¹¹¹In, ¹¹²In, ^(113m)In, ^(115m)In), technetium (⁹⁹Tc, ^(99m)Tc), thallium (²⁰¹Ti), gallium (⁶⁸Ga, ⁶⁷Ga), palladium (¹⁰³Pd), molybdenum (⁹⁹Mo), xenon (¹³³Xe), fluorine (¹⁸F), ¹⁵³Sm, ¹⁷⁷Lu, ¹⁵⁹Gd, ¹⁴⁹Pm, ¹⁴⁰La, ¹⁷⁵Yb, ¹⁶⁶Ho, ⁹⁰Y, ⁴⁷Sc, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁴²Pr, ¹⁰⁵Rh, and ⁹⁷Ru.

In specific embodiments, a polypeptide of the present invention or fragment or variant thereof is attached to macrocyclic chelators that associate with radiometal ions, including but not limited to, ¹⁷⁷Lu, ⁹⁰Y, ¹⁶⁶Ho, and ¹⁵³Sm, to polypeptides. In a preferred embodiment, the radiometal ion associated with the macrocyclic chelators is ¹¹¹In. In another preferred embodiment, the radiometal ion associated with the macrocyclic chelator is ⁹⁰Y. In specific embodiments, the macrocyclic chelator is 1,4,7,10-tetraazacyclododecane-N,N′,N″,N′″-tetraacetic acid (DOTA). In other specific embodiments, DOTA is attached to an antibody of the invention or fragment thereof via a linker molecule. Examples of linker molecules useful for conjugating DOTA to a polypeptide are commonly known in the art—see, for example, DeNardo et al., Clin Cancer Res. 4(10):2483-90 (1998); Peterson et al., Bioconjug. Chem. 10(4):553-7 (1999); and Zimmerman et al, Nucl. Med. Biol. 26(8):943-50 (1999); which are hereby incorporated by reference in their entirety.

As mentioned, the proteins of the invention may be modified by either natural processes, such as posttranslational processing, or by chemical modification techniques which are well known in the art. It will be appreciated that the same type of modification may be present in the same or varying degrees at several sites in a given polypeptide. Polypeptides of the invention may be branched, for example, as a result of ubiquitination, and they may be cyclic, with or without branching. Cyclic, branched, and branched cyclic polypeptides may result from posttranslation natural processes or may be made by synthetic methods. Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cysteine, formation of pyroglutamate, formylation, gamma-carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, pegylation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination. (See, for instance, PROTEINS—STRUCTURE AND MOLECULAR PROPERTIES, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, New York (1993); POSTTRANSLATIONAL COVALENT MODIFICATION OF PROTEINS, B. C. Johnson, Ed., Academic Press, New York, pgs. 1-12 (1983); Seifter et al., Meth. Enzymol. 182:626-646 (1990); Rattan et al., Ann. N.Y. Acad. Sci. 663:48-62 (1992)).

Also provided by the invention are chemically modified derivatives of the polypeptides of the invention which may provide additional advantages such as increased solubility, stability and circulating time of the polypeptide, or decreased immunogenicity (see U.S. Pat. No. 4,179,337). The chemical moieties for derivitization may be selected from water soluble polymers such as polyethylene glycol, ethylene glycol/propylene glycol copolymers, carboxymethylcellulose, dextran, polyvinyl alcohol and the like. The polypeptides may be modified at random positions within the molecule, or at predetermined positions within the molecule and may include one, two, three or more attached chemical moieties.

The polymer may be of any molecular weight, and may be branched or unbranched. For polyethylene glycol, the preferred molecular weight is between about 1 kDa and about 100 kDa (the term “about” indicating that in preparations of polyethylene glycol, some molecules will weigh more, some less, than the stated molecular weight) for ease in handling and manufacturing. Other sizes may be used, depending on the desired therapeutic profile (e.g., the duration of sustained release desired, the effects, if any on biological activity, the ease in handling, the degree or lack of antigenicity and other known effects of the polyethylene glycol to a therapeutic protein or analog). For example, the polyethylene glycol may have an average molecular weight of about 200, 500, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 6500, 7000, 7500, 8000, 8500, 9000, 9500, 10,000, 10,500, 11,000, 11,500, 12,000, 12,500, 13,000, 13,500, 14,000, 14,500, 15,000, 15,500, 16,000, 16,500, 17,000, 17,500, 18,000, 18,500, 19,000, 19,500, 20,000, 25,000, 30,000, 35,000, 40,000, 45,000, 50,000, 55,000, 60,000, 65,000, 70,000, 75,000, 80,000, 85,000, 90,000, 95,000, or 100,000 kDa.

As noted above, the polyethylene glycol may have a branched structure. Branched polyethylene glycols are described, for example, in U.S. Pat. No. 5,643,575; Morpurgo et al., Appl. Biochem. Biotechnol. 56:59-72 (1996); Vorobjev et al., Nucleosides Nucleotides 18:2745-2750 (1999); and Caliceti et al., Bioconjug. Chem. 10:638-646 (1999), the disclosures of each of which are incorporated herein by reference.

The polyethylene glycol molecules (or other chemical moieties) should be attached to the protein with consideration of effects on functional or antigenic domains of the protein. There are a number of attachment methods available to those skilled in the art, such as, for example, the method disclosed in EP 0 401 384 (coupling PEG to G-CSF), herein incorporated by reference; see also Malik et al., Exp. Hematol. 20:1028-1035 (1992), reporting pegylation of GM-CSF using tresyl chloride. For example, polyethylene glycol may be covalently bound through amino acid residues via a reactive group, such as a free amino or carboxyl group. Reactive groups are those to which an activated polyethylene glycol molecule may be bound. The amino acid residues having a free amino group may include lysine residues and the N-terminal amino acid residues; those having a free carboxyl group may include aspartic acid residues glutamic acid residues and the C-terminal amino acid residue. Sulfhydryl groups may also be used as a reactive group for attaching the polyethylene glycol molecules. Preferred for therapeutic purposes is attachment at an amino group, such as attachment at the N-terminus or lysine group.

As suggested above, polyethylene glycol may be attached to proteins via linkage to any of a number of amino acid residues. For example, polyethylene glycol can be linked to proteins via covalent bonds to lysine, histidine, aspartic acid, glutamic acid, or cysteine residues. One or more reaction chemistries may be employed to attach polyethylene glycol to specific amino acid residues (e.g., lysine, histidine, aspartic acid, glutamic acid, or cysteine) of the protein or to more than one type of amino acid residue (e.g., lysine, histidine, aspartic acid, glutamic acid, cysteine and combinations thereof) of the protein.

One may specifically desire proteins chemically modified at the N-terminus. Using polyethylene glycol as an illustration of the present composition, one may select from a variety of polyethylene glycol molecules (by molecular weight, branching, etc.), the proportion of polyethylene glycol molecules to protein (polypeptide) molecules in the reaction mix, the type of pegylation reaction to be performed, and the method of obtaining the selected N-terminally pegylated protein. The method of obtaining the N-terminally pegylated preparation (i.e., separating this moiety from other monopegylated moieties if necessary) may be by purification of the N-terminally pegylated material from a population of pegylated protein molecules. Selective proteins chemically modified at the N-terminus modification may be accomplished by reductive alkylation that exploits differential reactivity of different types of primary amino groups (lysine versus the N-terminal) available for derivatization in a particular protein. Under the appropriate reaction conditions, substantially selective derivatization of the protein at the N-terminus with a carbonyl group containing polymer is achieved.

As indicated above, pegylation of the proteins of the invention may be accomplished by any number of means. For example, polyethylene glycol may be attached to the protein either directly or by an intervening linker. Linkerless systems for attaching polyethylene glycol to proteins are described in Delgado et al., Crit. Rev. Thera. Drug Carrier Sys. 9:249-304 (1992); Francis et al., Intern. J. of Hematol. 68:1-18 (1998); U.S. Pat. No. 4,002,531; U.S. Pat. No. 5,349,052; WO 95/06058; and WO 98/32466, the disclosures of each of which are incorporated herein by reference.

One system for attaching polyethylene glycol directly to amino acid residues of proteins without an intervening linker employs tresylated MPEG, which is produced by the modification of monmethoxy polyethylene glycol (MPEG) using tresylchloride (ClSO₂CH₂CF₃). Upon reaction of protein with tresylated MPEG, polyethylene glycol is directly attached to amine groups of the protein. Thus, the invention includes protein-polyethylene glycol conjugates produced by reacting proteins of the invention with a polyethylene glycol molecule having a 2,2,2-trifluoreothane sulphonyl group.

Polyethylene glycol can also be attached to proteins using a number of different intervening linkers. For example, U.S. Pat. No. 5,612,460, the entire disclosure of which is incorporated herein by reference, discloses urethane linkers for connecting polyethylene glycol to proteins. Protein-polyethylene glycol conjugates wherein the polyethylene glycol is attached to the protein by a linker can also be produced by reaction of proteins with compounds such as MPEG-succinimidylsuccinate, MPEG activated with 1,1′-carbonyldiimidazole, MPEG-2,4,5-trichloropenylcarbonate, MPEG-p-nitrophenolcarbonate, and various MPEG-succinate derivatives. A number of additional polyethylene glycol derivatives and reaction chemistries for attaching polyethylene glycol to proteins are described in International Publication No. WO 98/32466, the entire disclosure of which is incorporated herein by reference. Pegylated protein products produced using the reaction chemistries set out herein are included within the scope of the invention.

The number of polyethylene glycol moieties attached to each protein of the invention (i.e., the degree of substitution) may also vary. For example, the pegylated proteins of the invention may be linked, on average, to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 17, 20, or more polyethylene glycol molecules. Similarly, the average degree of substitution within ranges such as 1-3, 2-4, 3-5, 4-6, 5-7, 6-8, 7-9, 8-10, 9-11, 10-12, 11-13, 12-14, 13-15, 14-16, 15-17, 16-18, 17-19, or 18-20 polyethylene glycol moieties per protein molecule. Methods for determining the degree of substitution are discussed, for example, in Delgado et al., Crit. Rev. Thera. Drug Carrier Sys. 9:249-304 (1992).

The polypeptides of the invention can be recovered and purified from chemical synthesis and recombinant cell cultures by standard methods which include, but are not limited to, ammonium sulfate or ethanol precipitation, acid extraction, anion or cation exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxylapatite chromatography and lectin chromatography. Most preferably, high performance liquid chromatography (“HPLC”) is employed for purification. Well known techniques for refolding protein may be employed to regenerate active conformation when the polypeptide is denatured during isolation and/or purification.

The polypeptides of the invention may be in monomers or multimers (i.e., dimers, trimers, tetramers and higher multimers). Accordingly, the present invention relates to monomers and multimers of the polypeptides of the invention, their preparation, and compositions (preferably, Therapeutics) containing them. In specific embodiments, the polypeptides of the invention are monomers, dimers, trimers or tetramers. In additional embodiments, the multimers of the invention are at least dimers, at least trimers, or at least tetramers.

Multimers encompassed by the invention may be homomers or heteromers. As used herein, the term homomer refers to a multimer containing only polypeptides corresponding to a protein of the invention (e.g., the amino acid sequence of SEQ ID NO:Y, an amino acid sequence encoded by SEQ ID NO:X or the complement of SEQ ID NO:X, the amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, and/or an amino acid sequence encoded by cDNA contained in ATCC™ Deposit No:Z (including fragments, variants, splice variants, and fusion proteins, corresponding to these as described herein)). These homomers may contain polypeptides having identical or different amino acid sequences. In a specific embodiment, a homomer of the invention is a multimer containing only polypeptides having an identical amino acid sequence. In another specific embodiment, a homomer of the invention is a multimer containing polypeptides having different amino acid sequences. In specific embodiments, the multimer of the invention is a homodimer (e.g., containing two polypeptides having identical or different amino acid sequences) or a homotrimer (e.g., containing three polypeptides having identical and/or different amino acid sequences). In additional embodiments, the homomeric multimer of the invention is at least a homodimer, at least a homotrimer, or at least a homotetramer.

As used herein, the term heteromer refers to a multimer containing one or more heterologous polypeptides (i.e., polypeptides of different proteins) in addition to the polypeptides of the invention. In a specific embodiment, the multimer of the invention is a heterodimer, a heterotrimer, or a heterotetramer. In additional embodiments, the heteromeric multimer of the invention is at least a heterodimer, at least a heterotrimer, or at least a heterotetramer.

Multimers of the invention may be the result of hydrophobic, hydrophilic, ionic and/or covalent associations and/or may be indirectly linked by, for example, liposome formation. Thus, in one embodiment, multimers of the invention, such as, for example, homodimers or homotrimers, are formed when polypeptides of the invention contact one another in solution. In another embodiment, heteromultimers of the invention, such as, for example, heterotrimers or heterotetramers, are formed when polypeptides of the invention contact antibodies to the polypeptides of the invention (including antibodies to the heterologous polypeptide sequence in a fusion protein of the invention) in solution. In other embodiments, multimers of the invention are formed by covalent associations with and/or between the polypeptides of the invention. Such covalent associations may involve one or more amino acid residues contained in the polypeptide sequence (e.g., that recited in SEQ ID NO:Y, encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, and/or encoded by the cDNA contained in ATCC™ Deposit No:Z). In one instance, the covalent associations are cross-linking between cysteine residues located within the polypeptide sequences which interact in the native (i.e., naturally occurring) polypeptide. In another instance, the covalent associations are the consequence of chemical or recombinant manipulation. Alternatively, such covalent associations may involve one or more amino acid residues contained in the heterologous polypeptide sequence in a fusion protein. In one example, covalent associations are between the heterologous sequence contained in a fusion protein of the invention (see, e.g., U.S. Pat. No. 5,478,925). In a specific example, the covalent associations are between the heterologous sequence contained in a Fc fusion protein of the invention (as described herein). In another specific example, covalent associations of fusion proteins of the invention are between heterologous polypeptide sequence from another protein that is capable of forming covalently associated multimers, such as for example, osteoprotegerin (see, e.g., International Publication NO: WO 98/49305, the contents of which are herein incorporated by reference in its entirety). In another embodiment, two or more polypeptides of the invention are joined through peptide linkers. Examples include those peptide linkers described in U.S. Pat. No. 5,073,627 (hereby incorporated by reference). Proteins comprising multiple polypeptides of the invention separated by peptide linkers may be produced using conventional recombinant DNA technology.

Another method for preparing multimer polypeptides of the invention involves use of polypeptides of the invention fused to a leucine zipper or isoleucine zipper polypeptide sequence. Leucine zipper and isoleucine zipper domains are polypeptides that promote multimerization of the proteins in which they are found. Leucine zippers were originally identified in several DNA-binding proteins (Landschulz et al., Science 240:1759, (1988)), and have since been found in a variety of different proteins. Among the known leucine zippers are naturally occurring peptides and derivatives thereof that dimerize or trimerize. Examples of leucine zipper domains suitable for producing soluble multimeric proteins of the invention are those described in PCT application WO 94/10308, hereby incorporated by reference. Recombinant fusion proteins comprising a polypeptide of the invention fused to a polypeptide sequence that dimerizes or trimerizes in solution are expressed in suitable host cells, and the resulting soluble multimeric fusion protein is recovered from the culture supernatant using techniques known in the art.

Trimeric polypeptides of the invention may offer the advantage of enhanced biological activity. Preferred leucine zipper moieties and isoleucine moieties are those that preferentially form trimers. One example is a leucine zipper derived from lung surfactant protein D (SPD), as described in Hoppe et al. (FEBS Letters 344:191, (1994)) and in U.S. patent application Ser. No. 08/446,922, hereby incorporated by reference. Other peptides derived from naturally occurring trimeric proteins may be employed in preparing trimeric polypeptides of the invention.

In another example, proteins of the invention are associated by interactions between Flag® polypeptide sequence contained in fusion proteins of the invention containing Flag® polypeptide sequence. In a further embodiment, proteins of the invention are associated by interactions between heterologous polypeptide sequence contained in Flag® fusion proteins of the invention and anti-Flag® antibody.

The multimers of the invention may be generated using chemical techniques known in the art. For example, polypeptides desired to be contained in the multimers of the invention may be chemically cross-linked using linker molecules and linker molecule length optimization techniques known in the art (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). Additionally, multimers of the invention may be generated using techniques known in the art to form one or more inter-molecule cross-links between the cysteine residues located within the sequence of the polypeptides desired to be contained in the multimer (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). Further, polypeptides of the invention may be routinely modified by the addition of cysteine or biotin to the C-terminus or N-terminus of the polypeptide and techniques known in the art may be applied to generate multimers containing one or more of these modified polypeptides (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). Additionally, techniques known in the art may be applied to generate liposomes containing the polypeptide components desired to be contained in the multimer of the invention (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety).

Alternatively, multimers of the invention may be generated using genetic engineering techniques known in the art. In one embodiment, polypeptides contained in multimers of the invention are produced recombinantly using fusion protein technology described herein or otherwise known in the art (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). In a specific embodiment, polynucleotides coding for a homodimer of the invention are generated by ligating a polynucleotide sequence encoding a polypeptide of the invention to a sequence encoding a linker polypeptide and then further to a synthetic polynucleotide encoding the translated product of the polypeptide in the reverse orientation from the original C-terminus to the N-terminus (lacking the leader sequence) (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). In another embodiment, recombinant techniques described herein or otherwise known in the art are applied to generate recombinant polypeptides of the invention which contain a transmembrane domain (or hydrophobic or signal peptide) and which can be incorporated by membrane reconstitution techniques into liposomes (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety).

Antibodies

Further polypeptides of the invention relate to antibodies and T-cell antigen receptors (TCR) which immunospecifically bind a polypeptide, polypeptide fragment, or variant of the invention (e.g., a polypeptide or fragment or variant of the amino acid sequence of SEQ ID NO:Y or a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z, and/or an epitope, of the present invention) as determined by immunoassays well known in the art for assaying specific antibody-antigen binding. Antibodies of the invention include, but are not limited to, polyclonal, monoclonal, multispecific, human, humanized or chimeric antibodies, single chain antibodies, Fab fragments, F(ab′) fragments, fragments produced by a Fab expression library, anti-idiotypic (anti-Id) antibodies (including, e.g., anti-Id antibodies to antibodies of the invention), intracellularly-made antibodies (i.e., intrabodies), and epitope-binding fragments of any of the above. The term “antibody,” as used herein, refers to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, i.e., molecules that contain an antigen binding site that immunospecifically binds an antigen. The immunoglobulin molecules of the invention can be of any type (e.g., IgG, IgE, IgM, IgD, IgA and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2) or subclass of immunoglobulin molecule. In preferred embodiments, the immunoglobulin molecules of the invention are IgG1. In other preferred embodiments, the immunoglobulin molecules of the invention are IgG4.

Most preferably the antibodies are human antigen-binding antibody fragments of the present invention and include, but are not limited to, Fab, Fab′ and F(ab′)2, Fd, single-chain Fvs (scFv), single-chain antibodies, disulfide-linked Fvs (sdFv) and fragments comprising either a VL or VH domain. Antigen-binding antibody fragments, including single-chain antibodies, may comprise the variable region(s) alone or in combination with the entirety or a portion of the following: hinge region, CH1, CH2, and CH3 domains. Also included in the invention are antigen-binding fragments also comprising any combination of variable region(s) with a hinge region, CH1, CH2, and CH3 domains. The antibodies of the invention may be from any animal origin including birds and mammals. Preferably, the antibodies are human, murine (e.g., mouse and rat), donkey, ship rabbit, goat, guinea pig, camel, horse, or chicken. As used herein, “human” antibodies include antibodies having the amino acid sequence of a human immunoglobulin and include antibodies isolated from human immunoglobulin libraries or from animals transgenic for one or more human immunoglobulin and that do not express endogenous immunoglobulins, as described infra and, for example in, U.S. Pat. No. 5,939,598 by Kucherlapati et al. The antibodies of the present invention may be monospecific, bispecific, trispecific or of greater multispecificity. Multispecific antibodies may be specific for different epitopes of a polypeptide of the present invention or may be specific for both a polypeptide of the present invention as well as for a heterologous epitope, such as a heterologous polypeptide or solid support material. See, e.g., PCT publications WO 93/17715; WO 92/08802; WO 91/00360; WO 92/05793; Tutt, et al., J. Immunol. 147:60-69 (1991); U.S. Pat. Nos. 4,474,893; 4,714,681; 4,925,648; 5,573,920; 5,601,819; Kostelny et al., J. Immunol. 148:1547-1553 (1992).

Antibodies of the present invention may be described or specified in terms of the epitope(s) or portion(s) of a polypeptide of the present invention which they recognize or specifically bind. The epitope(s) or polypeptide portion(s) may be specified as described herein, e.g., by N-terminal and C-terminal positions, or by size in contiguous amino acid residues, or listed in the Tables and Figures. Preferred epitopes of the invention include the predicted epitopes shown in Table 1B, as well as polynucleotides that encode these epitopes. Antibodies which specifically bind any epitope or polypeptide of the present invention may also be excluded. Therefore, the present invention includes antibodies that specifically bind polypeptides of the present invention, and allows for the exclusion of the same.

Antibodies of the present invention may also be described or specified in terms of their cross-reactivity. Antibodies that do not bind any other analog, ortholog, or homolog of a polypeptide of the present invention are included. Antibodies that bind polypeptides with at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 65%, at least 60%, at least 55%, and at least 50% identity (as calculated using methods known in the art and described herein) to a polypeptide of the present invention are also included in the present invention. In specific embodiments, antibodies of the present invention cross-react with murine, rat and/or rabbit homologs of human proteins and the corresponding epitopes thereof. Antibodies that do not bind polypeptides with less than 95%, less than 90%, less than 85%, less than 80%, less than 75%, less than 70%, less than 65%, less than 60%, less than 55%, and less than 50% identity (as calculated using methods known in the art and described herein) to a polypeptide of the present invention are also included in the present invention. In a specific embodiment, the above-described cross-reactivity is with respect to any single specific antigenic or immunogenic polypeptide, or combination(s) of 2, 3, 4, 5, or more of the specific antigenic and/or immunogenic polypeptides disclosed herein. Further included in the present invention are antibodies which bind polypeptides encoded by polynucleotides which hybridize to a polynucleotide of the present invention under stringent hybridization conditions (as described herein). Antibodies of the present invention may also be described or specified in terms of their binding affinity to a polypeptide of the invention. Preferred binding affinities include those with a dissociation constant or Kd less than 5×10⁻² M⁻², 10 M, 5×10⁻³ M, 10⁻³ M, 5×10⁻⁴ M, 10⁻⁴ M, 5×10⁻⁵ M, 10⁻⁵ M, 5×10⁻⁶ M, 10⁻⁶M, 5×10⁻⁷ M, 10⁷ M, 5×10⁻⁸ M, 10⁻⁸ M, 5×10⁻⁹ M, 10⁻⁹ M, 5×10⁻¹⁰ M, 10⁻¹⁰ M, 5×10⁻¹¹ M, 10⁻¹¹ M, 5×10⁻¹² M, 10⁻¹² M, 5×10⁻¹³ M, 10⁻¹³ M, 5×10⁻¹⁴ M, 10⁻¹⁴ M, 5×10⁻¹⁵ M, or 10⁻¹⁵ M.

The invention also provides antibodies that competitively inhibit binding of an antibody to an epitope of the invention as determined by any method known in the art for determining competitive binding, for example, the immunoassays described herein. In preferred embodiments, the antibody competitively inhibits binding to the epitope by at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 60%, or at least 50%.

Antibodies of the present invention may act as agonists or antagonists of the polypeptides of the present invention. For example, the present invention includes antibodies which disrupt the receptor/ligand interactions with the polypeptides of the invention either partially or fully. Preferably, antibodies of the present invention bind an antigenic epitope disclosed herein, or a portion thereof. The invention features both receptor-specific antibodies and ligand-specific antibodies. The invention also features receptor-specific antibodies which do not prevent ligand binding but prevent receptor activation. Receptor activation (i.e., signaling) may be determined by techniques described herein or otherwise known in the art. For example, receptor activation can be determined by detecting the phosphorylation (e.g., tyrosine or serine/threonine) of the receptor or its substrate by immunoprecipitation followed by western blot analysis (for example, as described supra). In specific embodiments, antibodies are provided that inhibit ligand activity or receptor activity by at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 60%, or at least 50% of the activity in absence of the antibody.

The invention also features receptor-specific antibodies which both prevent ligand binding and receptor activation as well as antibodies that recognize the receptor-ligand complex, and, preferably, do not specifically recognize the unbound receptor or the unbound ligand. Likewise, included in the invention are neutralizing antibodies which bind the ligand and prevent binding of the ligand to the receptor, as well as antibodies which bind the ligand, thereby preventing receptor activation, but do not prevent the ligand from binding the receptor. Further included in the invention are antibodies which activate the receptor. These antibodies may act as receptor agonists, i.e., potentiate or activate either all or a subset of the biological activities of the ligand-mediated receptor activation, for example, by inducing dimerization of the receptor. The antibodies may be specified as agonists, antagonists or inverse agonists for biological activities comprising the specific biological activities of the peptides of the invention disclosed herein. The above antibody agonists can be made using methods known in the art. See, e.g., PCT publication WO 96/40281; U.S. Pat. No. 5,811,097; Deng et al., Blood 92(6):1981-1988 (1998); Chen et al., Cancer Res. 58(16):3668-3678 (1998); Harrop et al., J. Immunol. 161(4):1786-1794 (1998); Zhu et al., Cancer Res. 58(15):3209-3214 (1998); Yoon et al., J. Immunol. 160(7):3170-3179 (1998); Prat et al., J. Cell. Sci. 111(Pt2):237-247 (1998); Pitard et al., J. Immunol. Methods 205 (2): 177-190 (1997); Liautard et al., Cytokine 9(4):233-241 (1997); Carlson et al., J. Biol. Chem. 272(17):11295-11301 (1997); Taryman et al., Neuron 14(4):755-762 (1995); Muller et al., Structure 6(9):1153-1167 (1998); Bartunek et al., Cytokine 8(1):14-20 (1996) (which are all incorporated by reference herein in their entireties).

Antibodies of the present invention may be used, for example, to purify, detect, and target the polypeptides of the present invention, including both in vitro and in vivo diagnostic and therapeutic methods. For example, the antibodies have utility in immunoassays for qualitatively and quantitatively measuring levels of the polypeptides of the present invention in biological samples. See, e.g., Harlow et al., Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988); incorporated by reference herein in its entirety.

As discussed in more detail below, the antibodies of the present invention may be used either alone or in combination with other compositions. The antibodies may further be recombinantly fused to a heterologous polypeptide at the N- or C-terminus or chemically conjugated (including covalent and non-covalent conjugations) to polypeptides or other compositions. For example, antibodies of the present invention may be recombinantly fused or conjugated to molecules useful as labels in detection assays and effector molecules such as heterologous polypeptides, drugs, radionuclides, or toxins. See, e.g., PCT publications WO 92/08495; WO 91/14438; WO 89/12624; U.S. Pat. No. 5,314,995; and EP 396,387; the disclosures of which are incorporated herein by reference in their entireties.

The antibodies of the invention include derivatives that are modified, i.e, by the covalent attachment of any type of molecule to the antibody such that covalent attachment does not prevent the antibody from generating an anti-idiotypic response. For example, but not by way of limitation, the antibody derivatives include antibodies that have been modified, e.g., by glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, linkage to a cellular ligand or other protein, etc. Any of numerous chemical modifications may be carried out by known techniques, including, but not limited to specific chemical cleavage, acetylation, formylation, metabolic synthesis of tunicamycin, etc. Additionally, the derivative may contain one or more non-classical amino acids.

The antibodies of the present invention may be generated by any suitable method known in the art. Polyclonal antibodies to an antigen-of-interest can be produced by various procedures well known in the art. For example, a polypeptide of the invention can be administered to various host animals including, but not limited to, rabbits, mice, rats, etc. to induce the production of sera containing polyclonal antibodies specific for the antigen. Various adjuvants may be used to increase the immunological response, depending on the host species, and include but are not limited to, Freund's (complete and incomplete), mineral gels such as aluminum hydroxide, surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, keyhole limpet hemocyanins, dinitrophenol, and potentially useful human adjuvants such as BCG (bacille Calmette-Guerin) and corynebacterium parvum. Such adjuvants are also well known in the art.

Monoclonal antibodies can be prepared using a wide variety of techniques known in the art including the use of hybridoma, recombinant, and phage display technologies, or a combination thereof. For example, monoclonal antibodies can be produced using hybridoma techniques including those known in the art and taught, for example, in Harlow et al., Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988); Hammerling, et al., in: Monoclonal Antibodies and T-Cell Hybridomas 563-681 (Elsevier, N.Y., 1981) (said references incorporated by reference in their entireties). The term “monoclonal antibody” as used herein is not limited to antibodies produced through hybridoma technology. The term “monoclonal antibody” refers to an antibody that is derived from a single clone, including any eukaryotic, prokaryotic, or phage clone, and not the method by which it is produced.

Methods for producing and screening for specific antibodies using hybridoma technology are routine and well known in the art and are discussed in detail in the Examples. In a non-limiting example, mice can be immunized with a polypeptide of the invention or a cell expressing such peptide. Once an immune response is detected, e.g., antibodies specific for the antigen are detected in the mouse serum, the mouse spleen is harvested and splenocytes isolated. The splenocytes are then fused by well known techniques to any suitable myeloma cells, for example cells from cell line SP20 available from the ATCC™. Hybridomas are selected and cloned by limited dilution. The hybridoma clones are then assayed by methods known in the art for cells that secrete antibodies capable of binding a polypeptide of the invention. Ascites fluid, which generally contains high levels of antibodies, can be generated by immunizing mice with positive hybridoma clones.

Accordingly, the present invention provides methods of generating monoclonal antibodies as well as antibodies produced by the method comprising culturing a hybridoma cell secreting an antibody of the invention wherein, preferably, the hybridoma is generated by fusing splenocytes isolated from a mouse immunized with an antigen of the invention with myeloma cells and then screening the hybridomas resulting from the fusion for hybridoma clones that secrete an antibody able to bind a polypeptide of the invention.

Another well known method for producing both polyclonal and monoclonal human B cell lines is transformation using Epstein Barr Virus (EBV). Protocols for generating EBV-transformed B cell lines are commonly known in the art, such as, for example, the protocol outlined in Chapter 7.22 of Current Protocols in Immunology, Coligan et al., Eds., 1994, John Wiley & Sons, NY, which is hereby incorporated in its entirety by reference. The source of B cells for transformation is commonly human peripheral blood, but B cells for transformation may also be derived from other sources including, but not limited to, lymph nodes, tonsil, spleen, tumor tissue, and infected tissues. Tissues are generally made into single cell suspensions prior to EBV transformation. Additionally, steps may be taken to either physically remove or inactivate T cells (e.g., by treatment with cyclosporin A) in B cell-containing samples, because T cells from individuals seropositive for anti-EBV antibodies can suppress B cell immortalization by EBV.

In general, the sample containing human B cells is innoculated with EBV, and cultured for 3-4 weeks. A typical source of EBV is the culture supernatant of the B95-8 cell line (ATCC™ #VR-1492). Physical signs of EBV transformation can generally be seen towards the end of the 3-4 week culture period. By phase-contrast microscopy, transformed cells may appear large, clear, hairy and tend to aggregate in tight clusters of cells. Initially, EBV lines are generally polyclonal. However, over prolonged periods of cell cultures, EBV lines may become monoclonal or polyclonal as a result of the selective outgrowth of particular B cell clones. Alternatively, polyclonal EBV transformed lines may be subcloned (e.g., by limiting dilution culture) or fused with a suitable fusion partner and plated at limiting dilution to obtain monoclonal B cell lines. Suitable fusion partners for EBV transformed cell lines include mouse myeloma cell lines (e.g., SP2/0, X63-Ag8.653), heteromyeloma cell lines (human×mouse; e.g, SPAM-8, SBC-H20, and CB-F7), and human cell lines (e.g., GM 1500, SKO-007, RPMI 8226, and KR-4). Thus, the present invention also provides a method of generating polyclonal or monoclonal human antibodies against polypeptides of the invention or fragments thereof, comprising EBV-transformation of human B cells.

Antibody fragments which recognize specific epitopes may be generated by known techniques. For example, Fab and F(ab′)2 fragments of the invention may be produced by proteolytic cleavage of immunoglobulin molecules, using enzymes such as papain (to produce Fab fragments) or pepsin (to produce F(ab′)2 fragments). F(ab′)2 fragments contain the variable region, the light chain constant region and the CH1 domain of the heavy chain.

For example, the antibodies of the present invention can also be generated using various phage display methods known in the art. In phage display methods, functional antibody domains are displayed on the surface of phage particles which carry the polynucleotide sequences encoding them. In a particular embodiment, such phage can be utilized to display antigen binding domains expressed from a repertoire or combinatorial antibody library (e.g., human or murine). Phage expressing an antigen binding domain that binds the antigen of interest can be selected or identified with antigen, e.g., using labeled antigen or antigen bound or captured to a solid surface or bead. Phage used in these methods are typically filamentous phage including fd and M13 binding domains expressed from phage with Fab, Fv or disulfide stabilized Fv antibody domains recombinantly fused to either the phage gene III or gene VIII protein. Examples of phage display methods that can be used to make the antibodies of the present invention include those disclosed in Brinkman et al., J. Immunol. Methods 182:41-50 (1995); Ames et al., J. Immunol. Methods 184:177-186 (1995); Kettleborough et al., Eur. J. Immunol. 24:952-958 (1994); Persic et al., Gene 187 9-18 (1997); Burton et al., Advances in Immunology 57:191-280 (1994); PCT application No. PCT/GB91/01134; PCT publications WO 90/02809; WO 91/10737; WO 92/01047; WO 92/18619; WO 93/11236; WO 95/15982; WO 95/20401; and U.S. Pat. Nos. 5,698,426; 5,223,409; 5,403,484; 5,580,717; 5,427,908; 5,750,753; 5,821,047; 5,571,698; 5,427,908; 5,516,637; 5,780,225; 5,658,727; 5,733,743 and 5,969,108; each of which is incorporated herein by reference in its entirety.

As described in the above references, after phage selection, the antibody coding regions from the phage can be isolated and used to generate whole antibodies, including human antibodies, or any other desired antigen binding fragment, and expressed in any desired host, including mammalian cells, insect cells, plant cells, yeast, and bacteria, e.g., as described in detail below. For example, techniques to recombinantly produce Fab, Fab′ and F(ab′)2 fragments can also be employed using methods known in the art such as those disclosed in PCT publication WO 92/22324; Mullinax et al., BioTechniques 12(6):864-869 (1992); and Sawai et al., AJRI 34:26-34 (1995); and Better et al., Science 240:1041-1043 (1988) (said references incorporated by reference in their entireties).

Examples of techniques which can be used to produce single-chain Fvs and antibodies include those described in U.S. Pat. Nos. 4,946,778 and 5,258,498; Huston et al., Methods in Enzymology 203:46-88 (1991); Shu et al., PNAS 90:7995-7999 (1993); and Skerra et al., Science 240:1038-1040 (1988). For some uses, including in vivo use of antibodies in humans and in vitro detection assays, it may be preferable to use chimeric, humanized, or human antibodies. A chimeric antibody is a molecule in which different portions of the antibody are derived from different animal species, such as antibodies having a variable region derived from a murine monoclonal antibody and a human immunoglobulin constant region. Methods for producing chimeric antibodies are known in the art. See e.g., Morrison, Science 229:1202 (1985); Oi et al., BioTechniques 4:214 (1986); Gillies et al., (1989) J. Immunol Methods 125:191-202; U.S. Pat. Nos. 5,807,715; 4,816,567; and 4,816,397, which are incorporated herein by reference in their entirety. Humanized antibodies are antibody molecules from non-human species antibody that binds the desired antigen having one or more complementarity determining regions (CDRs) from the non-human species and a framework region from a human immunoglobulin molecule. Often, framework residues in the human framework regions will be substituted with the corresponding residue from the CDR donor antibody to alter, preferably improve, antigen binding. These framework substitutions are identified by methods well known in the art, e.g., by modeling of the interactions of the CDR and framework residues to identify framework residues important for antigen binding and sequence comparison to identify unusual framework residues at particular positions. (See, e.g., Queen et al., U.S. Pat. No. 5,585,089; Riechmann et al., Nature 332:323 (1988), which are incorporated herein by reference in their entireties.) Antibodies can be humanized using a variety of techniques known in the art including, for example, CDR-grafting (EP 239,400; PCT publication WO 91/09967; U.S. Pat. Nos. 5,225,539; 5,530,101; and 5,585,089), veneering or resurfacing (EP 592,106; EP 519,596; Padlan, Molecular Immunology 28(4/5):489-498 (1991); Studnicka et al., Protein Engineering 7(6):805-814 (1994); Roguska. et al., PNAS 91:969-973 (1994)), and chain shuffling (U.S. Pat. No. 5,565,332).

Completely human antibodies are particularly desirable for therapeutic treatment of human patients. Human antibodies can be made by a variety of methods known in the art including phage display methods described above using antibody libraries derived from human immunoglobulin sequences. See also, U.S. Pat. Nos. 4,444,887 and 4,716,111; and PCT publications WO 98/46645, WO 98/50433, WO 98/24893, WO 98/16654, WO 96/34096, WO 96/33735, and WO 91/10741; each of which is incorporated herein by reference in its entirety.

Human antibodies can also be produced using transgenic mice which are incapable of expressing functional endogenous immunoglobulins, but which can express human immunoglobulin genes. For example, the human heavy and light chain immunoglobulin gene complexes may be introduced randomly or by homologous recombination into mouse embryonic stem cells. Alternatively, the human variable region, constant region, and diversity region may be introduced into mouse embryonic stem cells in addition to the human heavy and light chain genes. The mouse heavy and light chain immunoglobulin genes may be rendered non-functional separately or simultaneously with the introduction of human immunoglobulin loci by homologous recombination. In particular, homozygous deletion of the JH region prevents endogenous antibody production. The modified embryonic stem cells are expanded and microinjected into blastocysts to produce chimeric mice. The chimeric mice are then bred to produce homozygous offspring which express human antibodies. The transgenic mice are immunized in the normal fashion with a selected antigen, e.g., all or a portion of a polypeptide of the invention. Monoclonal antibodies directed against the antigen can be obtained from the immunized, transgenic mice using conventional hybridoma technology. The human immunoglobulin transgenes harbored by the transgenic mice rearrange during B cell differentiation, and subsequently undergo class switching and somatic mutation. Thus, using such a technique, it is possible to produce therapeutically useful IgG, IgA, IgM and IgE antibodies. For an overview of this technology for producing human antibodies, see Lonberg and Huszar, Int. Rev. Immunol 13:65-93 (1995). For a detailed discussion of this technology for producing human antibodies and human monoclonal antibodies and protocols for producing such antibodies, see, e.g., PCT publications WO 98/24893; WO 92/01047; WO 96/34096; WO 96/33735; European Patent No. 0 598 877; U.S. Pat. Nos. 5,413,923; 5,625,126; 5,633,425; 5,569,825; 5,661,016; 5,545,806; 5,814,318; 5,885,793; 5,916,771; 5,939,598; 6,075,181; and 6,114,598, which are incorporated by reference herein in their entirety. In addition, companies such as Abgenix, Inc. (Freemont, Calif.) and Genpharm (San Jose, Calif.) can be engaged to provide human antibodies directed against a selected antigen using technology similar to that described above.

Completely human antibodies which recognize a selected epitope can be generated using a technique referred to as “guided selection.” In this approach a selected non-human monoclonal antibody, e.g., a mouse antibody, is used to guide the selection of a completely human antibody recognizing the same epitope. (Jespers et al., Bio/technology 12:899-903 (1988)).

Further, antibodies to the polypeptides of the invention can, in turn, be utilized to generate anti-idiotype antibodies that “mimic” polypeptides of the invention using techniques well known to those skilled in the art. (See, e.g., Greenspan & Bona, FASEB J. 7(5):437-444; (1989) and Nissinoff, J. Immunol. 147(8):2429-2438 (1991)). For example, antibodies which bind to and competitively inhibit polypeptide multimerization and/or binding of a polypeptide of the invention to a ligand can be used to generate anti-idiotypes that “mimic” the polypeptide multimerization and/or binding domain and, as a consequence, bind to and neutralize polypeptide and/or its ligand. Such neutralizing anti-idiotypes or Fab fragments of such anti-idiotypes can be used in therapeutic regimens to neutralize polypeptide ligand(s)/receptor(s). For example, such anti-idiotypic antibodies can be used to bind a polypeptide of the invention and/or to bind its ligand(s)/receptor(s), and thereby block its biological activity. Alternatively, antibodies which bind to and enhance polypeptide multimerization and/or binding, and/or receptor/ligand multimerization, binding and/or signaling can be used to generate anti-idiotypes that function as agonists of a polypeptide of the invention and/or its ligand/receptor. Such agonistic anti-idiotypes or Fab fragments of such anti-idiotypes can be used in therapeutic regimens as agonists of the polypeptides of the invention or its ligand(s)/receptor(s). For example, such anti-idiotypic antibodies can be used to bind a polypeptide of the invention and/or to bind its ligand(s)/receptor(s), and thereby promote or enhance its biological activity.

Intrabodies of the invention can be produced using methods known in the art, such as those disclosed and reviewed in Chen et al., Hum. Gene Ther. 5:595-601 (1994); Marasco, W. A., Gene Ther. 4:11-15 (1997); Rondon and Marasco, Annu. Rev. Microbiol. 51:257-283 (1997); Proba et al., J. Mol. Biol. 275:245-253 (1998); Cohen et al., Oncogene 17:2445-2456 (1998); Ohage and Steipe, J. Mol. Biol. 291:1119-1128 (1999); Ohage et al., J. Mol. Biol. 291:1129-1134 (1999); Wirtz and Steipe, Protein Sci. 8:2245-2250 (1999); Zhu et al., J. Immunol. Methods 231:207-222 (1999); and references cited therein.

Polynucleotides Encoding Antibodies

The invention further provides polynucleotides comprising a nucleotide sequence encoding an antibody of the invention and fragments thereof. The invention also encompasses polynucleotides that hybridize under stringent or alternatively, under lower stringency hybridization conditions, e.g., as defined supra, to polynucleotides that encode an antibody, preferably, that specifically binds to a polypeptide of the invention, preferably, an antibody that binds to a polypeptide having the amino acid sequence of SEQ ID NO:Y, to a polypeptide encoded by a portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, and/or to a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

The polynucleotides may be obtained, and the nucleotide sequence of the polynucleotides determined, by any method known in the art. For example, if the nucleotide sequence of the antibody is known, a polynucleotide encoding the antibody may be assembled from chemically synthesized oligonucleotides (e.g., as described in Kutmeier et al., BioTechniques 17:242 (1994)), which, briefly, involves the synthesis of overlapping oligonucleotides containing portions of the sequence encoding the antibody, annealing and ligating of those oligonucleotides, and then amplification of the ligated oligonucleotides by PCR.

Alternatively, a polynucleotide encoding an antibody may be generated from nucleic acid from a suitable source. If a clone containing a nucleic acid encoding a particular antibody is not available, but the sequence of the antibody molecule is known, a nucleic acid encoding the immunoglobulin may be chemically synthesized or obtained from a suitable source (e.g., an antibody cDNA library, or a cDNA library generated from, or nucleic acid, preferably poly A+ RNA, isolated from, any tissue or cells expressing the antibody, such as hybridoma cells selected to express an antibody of the invention) by PCR amplification using synthetic primers hybridizable to the 3′ and 5′ ends of the sequence or by cloning using an oligonucleotide probe specific for the particular gene sequence to identify, e.g., a cDNA clone from a cDNA library that encodes the antibody. Amplified nucleic acids generated by PCR may then be cloned into replicable cloning vectors using any method well known in the art.

Once the nucleotide sequence and corresponding amino acid sequence of the antibody is determined, the nucleotide sequence of the antibody may be manipulated using methods well known in the art for the manipulation of nucleotide sequences, e.g., recombinant DNA techniques, site directed mutagenesis, PCR, etc. (see, for example, the techniques described in Sambrook et al., 1990, Molecular Cloning, A Laboratory Manual, 2d Ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y. and Ausubel et al., eds., 1998, Current Protocols in Molecular Biology, John Wiley & Sons, NY, which are both incorporated by reference herein in their entireties), to generate antibodies having a different amino acid sequence, for example to create amino acid substitutions, deletions, and/or insertions.

In a specific embodiment, the amino acid sequence of the heavy and/or light chain variable domains may be inspected to identify the sequences of the complementarity determining regions (CDRs) by methods that are well know in the art, e.g., by comparison to known amino acid sequences of other heavy and light chain variable regions to determine the regions of sequence hypervariability. Using routine recombinant DNA techniques, one or more of the CDRs may be inserted within framework regions, e.g., into human framework regions to humanize a non-human antibody, as described supra. The framework regions may be naturally occurring or consensus framework regions, and preferably human framework regions (see, e.g., Chothia et al., J. Mol. Biol. 278: 457-479 (1998) for a listing of human framework regions). Preferably, the polynucleotide generated by the combination of the framework regions and CDRs encodes an antibody that specifically binds a polypeptide of the invention. Preferably, as discussed supra, one or more amino acid substitutions may be made within the framework regions, and, preferably, the amino acid substitutions improve binding of the antibody to its antigen. Additionally, such methods may be used to make amino acid substitutions or deletions of one or more variable region cysteine residues participating in an intrachain disulfide bond to generate antibody molecules lacking one or more intrachain disulfide bonds. Other alterations to the polynucleotide are encompassed by the present invention and within the skill of the art.

In addition, techniques developed for the production of “chimeric antibodies” (Morrison et al., Proc. Natl. Acad. Sci. 81:851-855 (1984); Neuberger et al., Nature 312:604-608 (1984); Takeda et al., Nature 314:452-454 (1985)) by splicing genes from a mouse antibody molecule of appropriate antigen specificity together with genes from a human antibody molecule of appropriate biological activity can be used. As described supra, a chimeric antibody is a molecule in which different portions are derived from different animal species, such as those having a variable region derived from a murine mAb and a human immunoglobulin constant region, e.g., humanized antibodies.

Alternatively, techniques described for the production of single chain antibodies (U.S. Pat. No. 4,946,778; Bird, Science 242:423-42 (1988); Huston et al., Proc. Natl. Acad. Sci. USA 85:5879-5883 (1988); and Ward et al., Nature 334:544-54 (1989)) can be adapted to produce single chain antibodies. Single chain antibodies are formed by linking the heavy and light chain fragments of the Fv region via an amino acid bridge, resulting in a single chain polypeptide. Techniques for the assembly of functional Fv fragments in E. coli may also be used (Skerra et al., Science 242:1038-1041 (1988)).

Methods of Producing Antibodies

The antibodies of the invention can be produced by any method known in the art for the synthesis of antibodies, in particular, by chemical synthesis or preferably, by recombinant expression techniques. Methods of producing antibodies include, but are not limited to, hybridoma technology, EBV transformation, and other methods discussed herein as well as through the use recombinant DNA technology, as discussed below.

Recombinant expression of an antibody of the invention, or fragment, derivative or analog thereof, (e.g., a heavy or light chain of an antibody of the invention or a single chain antibody of the invention), requires construction of an expression vector containing a polynucleotide that encodes the antibody. Once a polynucleotide encoding an antibody molecule or a heavy or light chain of an antibody, or portion thereof (preferably containing the heavy or light chain variable domain), of the invention has been obtained, the vector for the production of the antibody molecule may be produced by recombinant DNA technology using techniques well known in the art. Thus, methods for preparing a protein by expressing a polynucleotide containing an antibody encoding nucleotide sequence are described herein. Methods which are well known to those skilled in the art can be used to construct expression vectors containing antibody coding sequences and appropriate transcriptional and translational control signals. These methods include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination. The invention, thus, provides replicable vectors comprising a nucleotide sequence encoding an antibody molecule of the invention, or a heavy or light chain thereof, or a heavy or light chain variable domain, operably linked to a promoter. Such vectors may include the nucleotide sequence encoding the constant region of the antibody molecule (see, e.g., PCT Publication WO 86/05807; PCT Publication WO 89/01036; and U.S. Pat. No. 5,122,464) and the variable domain of the antibody may be cloned into such a vector for expression of the entire heavy or light chain.

The expression vector is transferred to a host cell by conventional techniques and the transfected cells are then cultured by conventional techniques to produce an antibody of the invention. Thus, the invention includes host cells containing a polynucleotide encoding an antibody of the invention, or a heavy or light chain thereof, or a single chain antibody of the invention, operably linked to a heterologous promoter. In preferred embodiments for the expression of double-chained antibodies, vectors encoding both the heavy and light chains may be co-expressed in the host cell for expression of the entire immunoglobulin molecule, as detailed below.

A variety of host-expression vector systems may be utilized to express the antibody molecules of the invention. Such host-expression systems represent vehicles by which the coding sequences of interest may be produced and subsequently purified, but also represent cells which may, when transformed or transfected with the appropriate nucleotide coding sequences, express an antibody molecule of the invention in situ. These include but are not limited to microorganisms such as bacteria (e.g., E. coli, B. subtilis) transformed with recombinant bacteriophage DNA, plasmid DNA or cosmid DNA expression vectors containing antibody coding sequences; yeast (e.g., Saccharomyces, Pichia) transformed with recombinant yeast expression vectors containing antibody coding sequences; insect cell systems infected with recombinant virus expression vectors (e.g., baculovirus) containing antibody coding sequences; plant cell systems infected with recombinant virus expression vectors (e.g., cauliflower mosaic virus, CaMV; tobacco mosaic virus, TMV) or transformed with recombinant plasmid expression vectors (e.g., Ti plasmid) containing antibody coding sequences; or mammalian cell systems (e.g., COS, CHO, BHK, 293, 3T3 cells) harboring recombinant expression constructs containing promoters derived from the genome of mammalian cells (e.g., metallothionein promoter) or from mammalian viruses (e.g., the adenovirus late promoter; the vaccinia virus 7.5K promoter). Preferably, bacterial cells such as Escherichia coli, and more preferably, eukaryotic cells, especially for the expression of whole recombinant antibody molecule, are used for the expression of a recombinant antibody molecule. For example, mammalian cells such as Chinese hamster ovary cells (CHO), in conjunction with a vector such as the major intermediate early gene promoter element from human cytomegalovirus is an effective expression system for antibodies (Foecking et al., Gene 45:101 (1986); Cockett et al., Bio/Technology 8:2 (1990)).

In bacterial systems, a number of expression vectors may be advantageously selected depending upon the use intended for the antibody molecule being expressed. For example, when a large quantity of such a protein is to be produced, for the generation of pharmaceutical compositions of an antibody molecule, vectors which direct the expression of high levels of fusion protein products that are readily purified may be desirable. Such vectors include, but are not limited, to the E. coli expression vector pUR278 (Ruther et al., EMBO J. 2:1791 (1983)), in which the antibody coding sequence may be ligated individually into the vector in frame with the lac Z coding region so that a fusion protein is produced; pIN vectors (Inouye & Inouye, Nucleic Acids Res. 13:3101-3109 (1985); Van Heeke & Schuster, J. Biol. Chem. 24:5503-5509 (1989)); and the like. pGEX vectors may also be used to express foreign polypeptides as fusion proteins with glutathione 5-transferase (GST). In general, such fusion proteins are soluble and can easily be purified from lysed cells by adsorption and binding to matrix glutathione-agarose beads followed by elution in the presence of free glutathione. The pGEX vectors are designed to include thrombin or factor Xa protease cleavage sites so that the cloned target gene product can be released from the GST moiety.

In an insect system, Autographa californica nuclear polyhedrosis virus (AcNPV) is used as a vector to express foreign genes. The virus grows in Spodoptera frugiperda cells. The antibody coding sequence may be cloned individually into non-essential regions (for example the polyhedrin gene) of the virus and placed under control of an AcNPV promoter (for example the polyhedrin promoter).

In mammalian host cells, a number of viral-based expression systems may be utilized. In cases where an adenovirus is used as an expression vector, the antibody coding sequence of interest may be ligated to an adenovirus transcription/translation control complex, e.g., the late promoter and tripartite leader sequence. This chimeric gene may then be inserted in the adenovirus genome by in vitro or in vivo recombination. Insertion in a non-essential region of the viral genome (e.g., region E1 or E3) will result in a recombinant virus that is viable and capable of expressing the antibody molecule in infected hosts. (e.g., see Logan & Shenk, Proc. Natl. Acad. Sci. USA 81:355-359 (1984)). Specific initiation signals may also be required for efficient translation of inserted antibody coding sequences. These signals include the ATG initiation codon and adjacent sequences. Furthermore, the initiation codon must be in phase with the reading frame of the desired coding sequence to ensure translation of the entire insert. These exogenous translational control signals and initiation codons can be of a variety of origins, both natural and synthetic. The efficiency of expression may be enhanced by the inclusion of appropriate transcription enhancer elements, transcription terminators, etc. (see Bittner et al., Methods in Enzymol. 153:51-544 (1987)).

In addition, a host cell strain may be chosen which modulates the expression of the inserted sequences, or modifies and processes the gene product in the specific fashion desired. Such modifications (e.g., glycosylation) and processing (e.g., cleavage) of protein products may be important for the function of the protein. Different host cells have characteristic and specific mechanisms for the post-translational processing and modification of proteins and gene products. Appropriate cell lines or host systems can be chosen to ensure the correct modification and processing of the foreign protein expressed. To this end, eukaryotic host cells which possess the cellular machinery for proper processing of the primary transcript, glycosylation, and phosphorylation of the gene product may be used. Such mammalian host cells include but are not limited to CHO, VERY, BHK, Hela, COS, MDCK, 293, 3T3, WI38, and in particular, breast cancer cell lines such as, for example, BT483, Hs578T, HTB2, BT20 and T47D, and normal mammary gland cell line such as, for example, CRL7030 and Hs578Bst.

For long-term, high-yield production of recombinant proteins, stable expression is preferred. For example, cell lines which stably express the antibody molecule may be engineered. Rather than using expression vectors which contain viral origins of replication, host cells can be transformed with DNA controlled by appropriate expression control elements (e.g., promoter, enhancer, sequences, transcription terminators, polyadenylation sites, etc.), and a selectable marker. Following the introduction of the foreign DNA, engineered cells may be allowed to grow for 1-2 days in an enriched media, and then are switched to a selective media. The selectable marker in the recombinant plasmid confers resistance to the selection and allows cells to stably integrate the plasmid into their chromosomes and grow to form foci that in turn can be cloned and expanded into cell lines. This method may advantageously be used to engineer cell lines which express the antibody molecule. Such engineered cell lines may be particularly useful in screening and evaluation of compounds that interact directly or indirectly with the antibody molecule.

A number of selection systems may be used, including but not limited to the herpes simplex virus thymidine kinase (Wigler et al., Cell 11:223 (1977)), hypoxanthine-guanine phosphoribosyltransferase (Szybalska & Szybalski, Proc. Natl. Acad. Sci. USA 48:202 (1992)), and adenine phosphoribosyltransferase (Lowy et al., Cell 22:817 (1980)) genes can be employed in tk−, hgprt− or aprt− cells, respectively. Also, antimetabolite resistance can be used as the basis of selection for the following genes: dhfr, which confers resistance to methotrexate (Wigler et al., Natl. Acad. Sci. USA 77:357 (1980); O'Hare et al., Proc. Natl. Acad. Sci. USA 78:1527 (1981)); gpt, which confers resistance to mycophenolic acid (Mulligan & Berg, Proc. Natl. Acad. Sci. USA 78:2072 (1981)); neo, which confers resistance to the aminoglycoside G-418 Clinical Pharmacy 12:488-505; Wu and Wu, Biotherapy 3:87-95 (1991); Tolstoshev, Ann. Rev. Pharmacol. Toxicol. 32:573-596 (1993); Mulligan, Science 260:926-932 (1993); and Morgan and Anderson, Ann. Rev. Biochem. 62:191-217 (1993); May, 1993, TIB TECH 11(5):155-215 (1993)); and hygro, which confers resistance to hygromycin (Santerre et al., Gene 30:147 (1984)). Methods commonly known in the art of recombinant DNA technology may be routinely applied to select the desired recombinant clone, and such methods are described, for example, in Ausubel et al. (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993); Kriegler, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990); and in Chapters 12 and 13, Dracopoli et al. (eds), Current Protocols in Human Genetics, John Wiley & Sons, NY (1994); Colberre-Garapin et al., J. Mol. Biol. 150:1 (1981), which are incorporated by reference herein in their entireties.

The expression levels of an antibody molecule can be increased by vector amplification (for a review, see Bebbington and Hentschel, The use of vectors based on gene amplification for the expression of cloned genes in mammalian cells in DNA cloning, Vol. 3. (Academic Press, New York, 1987)). When a marker in the vector system expressing antibody is amplifiable, increase in the level of inhibitor present in culture of host cell will increase the number of copies of the marker gene. Since the amplified region is associated with the antibody gene, production of the antibody will also increase (Crouse et al., Mol. Cell. Biol. 3:257 (1983)).

Vectors which use glutamine synthase (GS) or DHFR as the selectable markers can be amplified in the presence of the drugs methionine sulphoximine or methotrexate, respectively. An advantage of glutamine synthase based vectors are the availability of cell lines (e.g., the murine myeloma cell line, NS0) which are glutamine synthase negative. Glutamine synthase expression systems can also function in glutamine synthase expressing cells (e.g. Chinese Hamster Ovary (CHO) cells) by providing additional inhibitor to prevent the functioning of the endogenous gene. A glutamine synthase expression system and components thereof are detailed in PCT publications: WO87/04462; WO86/05807; WO89/01036; WO89/10404; and WO91/06657 which are incorporated in their entireties by reference herein. Additionally, glutamine synthase expression vectors that may be used according to the present invention are commercially available from suppliers, including, for example Lonza Biologics, Inc. (Portsmouth, N.H.). Expression and production of monoclonal antibodies using a GS expression system in murine myeloma cells is described in Bebbington et al., Bio/technology 10:169 (1992) and in Biblia and Robinson Biotechnol. Prog. 11:1 (1995) which are incorporated in their entireties by reference herein.

The host cell may be co-transfected with two expression vectors of the invention, the first vector encoding a heavy chain derived polypeptide and the second vector encoding a light chain derived polypeptide. The two vectors may contain identical selectable markers which enable equal expression of heavy and light chain polypeptides. Alternatively, a single vector may be used which encodes, and is capable of expressing, both heavy and light chain polypeptides. In such situations, the light chain should be placed before the heavy chain to avoid an excess of toxic free heavy chain (Proudfoot, Nature 322:52 (1986); Kohler, Proc. Natl. Acad. Sci. USA 77:2197 (1980)). The coding sequences for the heavy and light chains may comprise cDNA or genomic DNA.

Once an antibody molecule of the invention has been produced by an animal, chemically synthesized, or recombinantly expressed, it may be purified by any method known in the art for purification of an immunoglobulin molecule, for example, by chromatography (e.g., ion exchange, affinity, particularly by affinity for the specific antigen after Protein A, and sizing column chromatography), centrifugation, differential solubility, or by any other standard technique for the purification of proteins. In addition, the antibodies of the present invention or fragments thereof can be fused to heterologous polypeptide sequences described herein or otherwise known in the art, to facilitate purification.

The present invention encompasses antibodies recombinantly fused or chemically conjugated (including both covalently and non-covalently conjugations) to a polypeptide (or portion thereof, preferably at least 10, 20, 30, 40, 50, 60, 70, 80, 90 or 100 amino acids of the polypeptide) of the present invention to generate fusion proteins. The fusion does not necessarily need to be direct, but may occur through linker sequences. The antibodies may be specific for antigens other than polypeptides (or portion thereof, preferably at least 10, 20, 30, 40, 50, 60, 70, 80, 90 or 100 amino acids of the polypeptide) of the present invention. For example, antibodies may be used to target the polypeptides of the present invention to particular cell types, either in vitro or in vivo, by fusing or conjugating the polypeptides of the present invention to antibodies specific for particular cell surface receptors. Antibodies fused or conjugated to the polypeptides of the present invention may also be used in in vitro immunoassays and purification methods using methods known in the art. See e.g., Harbor et al., supra, and PCT publication WO 93/21232; EP 439,095; Naramura et al., Immunol Lett. 39:91-99 (1994); U.S. Pat. No. 5,474,981; Gillies et al., PNAS 89:1428-1432 (1992); Fell et al., J. Immunol. 146:2446-2452 (1991), which are incorporated by reference in their entireties.

The present invention further includes compositions comprising the polypeptides of the present invention fused or conjugated to antibody domains other than the variable regions. For example, the polypeptides of the present invention may be fused or conjugated to an antibody Fc region, or portion thereof. The antibody portion fused to a polypeptide of the present invention may comprise the constant region, hinge region, CH1 domain, CH2 domain, and CH3 domain or any combination of whole domains or portions thereof. The polypeptides may also be fused or conjugated to the above antibody portions to form multimers. For example, Fc portions fused to the polypeptides of the present invention can form dimers through disulfide bonding between the Fc portions. Higher multimeric forms can be made by fusing the polypeptides to portions of IgA and IgM. Methods for fusing or conjugating the polypeptides of the present invention to antibody portions are known in the art. See, e.g., U.S. Pat. Nos. 5,336,603; 5,622,929; 5,359,046; 5,349,053; 5,447,851; 5,112,946; EP 307,434; EP 367,166; PCT publications WO 96/04388; WO 91/06570; Ashkenazi et al., Proc. Natl. Acad. Sci. USA 88:10535-10539 (1991); Zheng et al., J. Immunol. 154:5590-5600 (1995); and Vil et al., Proc. Natl. Acad. Sci. USA 89:11337-11341 (1992) (said references incorporated by reference in their entireties).

As discussed, supra, the polypeptides corresponding to a polypeptide, polypeptide fragment, or a variant of SEQ ID NO:Y may be fused or conjugated to the above antibody portions to increase the in vivo half life of the polypeptides or for use in immunoassays using methods known in the art. Further, the polypeptides corresponding to SEQ ID NO:Y may be fused or conjugated to the above antibody portions to facilitate purification. One reported example describes chimeric proteins consisting of the first two domains of the human CD4-polypeptide and various domains of the constant regions of the heavy or light chains of mammalian immunoglobulins. See EP 394,827; and Traunecker et al., Nature 331:84-86 (1988). The polypeptides of the present invention fused or conjugated to an antibody having disulfide-linked dimeric structures (due to the IgG) may also be more efficient in binding and neutralizing other molecules, than the monomeric secreted protein or protein fragment alone. See, for example, Fountoulakis et al., J. Biochem. 270:3958-3964 (1995). In many cases, the Fc part in a fusion protein is beneficial in therapy and diagnosis, and thus can result in, for example, improved pharmacokinetic properties. See, for example, EP A 232,262. Alternatively, deleting the Fc part after the fusion protein has been expressed, detected, and purified, would be desired. For example, the Fc portion may hinder therapy and diagnosis if the fusion protein is used as an antigen for immunizations. In drug discovery, for example, human proteins, such as hIL-5, have been fused with Fc portions for the purpose of high-throughput screening assays to identify antagonists of hIL-5. (See, Bennett et al., J. Molecular Recognition 8:52-58 (1995); Johanson et al., J. Biol. Chem. 270:9459-9471 (1995)).

Moreover, the antibodies or fragments thereof of the present invention can be fused to marker sequences, such as a peptide to facilitate purification. In preferred embodiments, the marker amino acid sequence is a hexa-histidine peptide, such as the tag provided in a pQE vector (QIAGEN, Inc., 9259 Eton Avenue, Chatsworth, Calif., 91311), among others, many of which are commercially available. As described in Gentz et al., Proc. Natl. Acad. Sci. USA 86:821-824 (1989), for instance, hexa-histidine provides for convenient purification of the fusion protein. Other peptide tags useful for purification include, but are not limited to, the “HA” tag, which corresponds to an epitope derived from the influenza hemagglutinin protein (Wilson et al., Cell 37:767 (1984)) and the “flag” tag.

The present invention further encompasses antibodies or fragments thereof conjugated to a diagnostic or therapeutic agent. The antibodies can be used diagnostically to, for example, monitor the development or progression of a tumor as part of a clinical testing procedure to, e.g., determine the efficacy of a given treatment regimen. Detection can be facilitated by coupling the antibody to a detectable substance. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials, bioluminescent materials, radioactive materials, positron emitting metals using various positron emission tomographies, and nonradioactive paramagnetic metal ions. The detectable substance may be coupled or conjugated either directly to the antibody (or fragment thereof) or indirectly, through an intermediate (such as, for example, a linker known in the art) using techniques known in the art. See, for example, U.S. Pat. No. 4,741,900 for metal ions which can be conjugated to antibodies for use as diagnostics according to the present invention. Examples of suitable enzymes include horseradish peroxidase, alkaline phosphatase, beta-galactosidase, or acetylcholinesterase; examples of suitable prosthetic group complexes include streptavidin/biotin and avidin biotin; examples of suitable fluorescent materials include umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; an example of a luminescent material includes luminol; examples of bioluminescent materials include luciferase, luciferin, and aequorin; and examples of suitable radioactive material include 125I, 131I, 111In or 99Tc.

Further, an antibody or fragment thereof may be conjugated to a therapeutic moiety such as a cytotoxin, e.g., a cytostatic or cytocidal agent, a therapeutic agent or a radioactive metal ion, e.g., alpha-emitters such as, for example, 213Bi. A cytotoxin or cytotoxic agent includes any agent that is detrimental to cells. Examples include paclitaxol, cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicin, doxorubicin, daunorubicin, dihydroxy anthracin dione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, and puromycin and analogs or homologs thereof. Therapeutic agents include, but are not limited to, antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, 5-fluorouracil decarbazine), alkylating agents (e.g., mechlorethamine, thioepa chlorambucil, melphalan, carmustine (BSNU) and lomustine (CCNU), cyclothosphamide, busulfan, dibromomannitol, streptozotocin, mitomycin C, and cis-dichlorodiamine platinum (II) (DDP) cisplatin), anthracyclines (e.g., daunorubicin (formerly daunomycin) and doxorubicin), antibiotics (e.g., dactinomycin (formerly actinomycin), bleomycin, mithramycin, and anthramycin (AMC)), and anti-mitotic agents (e.g., vincristine and vinblastine).

The conjugates of the invention can be used for modifying a given biological response, the therapeutic agent or drug moiety is not to be construed as limited to classical chemical therapeutic agents. For example, the drug moiety may be a protein or polypeptide possessing a desired biological activity. Such proteins may include, for example, a toxin such as abrin, ricin A, pseudomonas exotoxin, or diphtheria toxin; a protein such as tumor necrosis factor, a-interferon, β-interferon, nerve growth factor, platelet derived growth factor, tissue plasminogen activator, an apoptotic agent, e.g., TNF-alpha, TNF-beta, AIM I (See, International Publication No. WO 97/33899), AIM II (See, International Publication No. WO 97/34911), Fas Ligand (Takahashi et al., Int. Immunol., 6:1567-1574 (1994)), VEGI (See, International Publication No. WO 99/23105), a thrombotic agent or an anti-angiogenic agent, e.g., angiostatin or endostatin; or, biological response modifiers such as, for example, lymphokines, interleukin-1 (“IL-1”), interleukin-2 (“IL-2”), interleukin-6 (“IL-6”), granulocyte macrophage colony stimulating factor (“GM-CSF”), granulocyte colony stimulating factor (“G-CSF”), or other growth factors.

Antibodies may also be attached to solid supports, which are particularly useful for immunoassays or purification of the target antigen. Such solid supports include, but are not limited to, glass, cellulose, polyacrylamide, nylon, polystyrene, polyvinyl chloride or polypropylene.

Techniques for conjugating such therapeutic moiety to antibodies are well known. See, for example, Amon et al., “Monoclonal Antibodies For Immunotargeting Of Drugs In Cancer Therapy”, in Monoclonal Antibodies And Cancer Therapy, Reisfeld et al. (eds.), pp. 243-56 (Alan R. Liss, Inc. 1985); Hellstrom et al., “Antibodies For Drug Delivery”, in Controlled Drug Delivery (2nd Ed.), Robinson et al. (eds.), pp. 623-53 (Marcel Dekker, Inc. 1987); Thorpe, “Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review”, in Monoclonal Antibodies '84: Biological And Clinical Applications, Pinchera et al. (eds.), pp. 475-506 (1985); “Analysis, Results, And Future Prospective Of The Therapeutic Use Of Radiolabeled Antibody In Cancer Therapy”, in Monoclonal Antibodies For Cancer Detection And Therapy, Baldwin et al. (eds.), pp. 303-16 (Academic Press 1985), and Thorpe et al., “The Preparation And Cytotoxic Properties Of Antibody-Toxin Conjugates”, Immunol Rev. 62:119-58 (1982).

Alternatively, an antibody can be conjugated to a second antibody to form an antibody heteroconjugate as described by Segal in U.S. Pat. No. 4,676,980, which is incorporated herein by reference in its entirety.

An antibody, with or without a therapeutic moiety conjugated to it, administered alone or in combination with cytotoxic factor(s) and/or cytokine(s) can be used as a therapeutic.

Immunophenotyping

The antibodies of the invention may be utilized for immunophenotyping of cell lines and biological samples. Translation products of the gene of the present invention may be useful as cell-specific markers, or more specifically as cellular markers that are differentially expressed at various stages of differentiation and/or maturation of particular cell types. Monoclonal antibodies directed against a specific epitope, or combination of epitopes, will allow for the screening of cellular populations expressing the marker. Various techniques can be utilized using monoclonal antibodies to screen for cellular populations expressing the marker(s), and include magnetic separation using antibody-coated magnetic beads, “panning” with antibody attached to a solid matrix (i.e., plate), and flow cytometry (See, e.g., U.S. Pat. No. 5,985,660; and Morrison et al., Cell, 96:737-49 (1999)).

These techniques allow for the screening of particular populations of cells, such as might be found with hematological malignancies (i.e. minimal residual disease (MRD) in acute leukemic patients) and “non-self” cells in transplantations to prevent Graft-versus-Host Disease (GVHD). Alternatively, these techniques allow for the screening of hematopoietic stem and progenitor cells capable of undergoing proliferation and/or differentiation, as might be found in human umbilical cord blood.

Assays For Antibody Binding

The antibodies of the invention may be assayed for immunospecific binding by any method known in the art. The immunoassays which can be used include but are not limited to competitive and non-competitive assay systems using techniques such as western blots, radioimmunoassays, ELISA (enzyme linked immunosorbent assay), “sandwich” immunoassays, immunoprecipitation assays, precipitin reactions, gel diffusion precipitin reactions, immunodiffusion assays, agglutination assays, complement-fixation assays, immunoradiometric assays, fluorescent immunoassays, and protein A immunoassays, to name but a few. Such assays are routine and well known in the art (see, e.g., Ausubel et al, eds, 1994, Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, which is incorporated by reference herein in its entirety). Exemplary immunoassays are described briefly below (but are not intended by way of limitation).

Immunoprecipitation protocols generally comprise lysing a population of cells in a lysis buffer such as RIPA buffer (1% NP-40 or Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 0.15 M NaCl, 0.01 M sodium phosphate at pH 7.2, 1% Trasylol) supplemented with protein phosphatase and/or protease inhibitors (e.g., EDTA, PMSF, aprotinin, sodium vanadate), adding the antibody of interest to the cell lysate, incubating for a period of time (e.g., 1-4 hours) at 4° C., adding protein A and/or protein G sepharose beads to the cell lysate, incubating for about an hour or more at 4° C., washing the beads in lysis buffer and resuspending the beads in SDS/sample buffer. The ability of the antibody of interest to immunoprecipitate a particular antigen can be assessed by, e.g., western blot analysis. One of skill in the art would be knowledgeable as to the parameters that can be modified to increase the binding of the antibody to an antigen and decrease the background (e.g., pre-clearing the cell lysate with sepharose beads). For further discussion regarding immunoprecipitation protocols see, e.g., Ausubel et al., eds., (1994), Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, section 10.16.1.

Western blot analysis generally comprises preparing protein samples, electrophoresis of the protein samples in a polyacrylamide gel (e.g., 8%-20% SDS-PAGE depending on the molecular weight of the antigen), transferring the protein sample from the polyacrylamide gel to a membrane such as nitrocellulose, PVDF or nylon, blocking the membrane in blocking solution (e.g., PBS with 3% BSA or non-fat milk), washing the membrane in washing buffer (e.g., PBS-Tween 20), blocking the membrane with primary antibody (the antibody of interest) diluted in blocking buffer, washing the membrane in washing buffer, blocking the membrane with a secondary antibody (which recognizes the primary antibody, e.g., an anti-human antibody) conjugated to an enzymatic substrate (e.g., horseradish peroxidase or alkaline phosphatase) or radioactive molecule (e.g., 32P or 125I) diluted in blocking buffer, washing the membrane in wash buffer, and detecting the presence of the antigen. One of skill in the art would be knowledgeable as to the parameters that can be modified to increase the signal detected and to reduce the background noise. For further discussion regarding western blot protocols see, e.g., Ausubel et al, eds, (1994), Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, section 10.8.1.

ELISAs comprise preparing antigen, coating the well of a 96 well microtiter plate with the antigen, adding the antibody of interest conjugated to a detectable compound such as an enzymatic substrate (e.g., horseradish peroxidase or alkaline phosphatase) to the well and incubating for a period of time, and detecting the presence of the antigen. In ELISAs the antibody of interest does not have to be conjugated to a detectable compound; instead, a second antibody (which recognizes the antibody of interest) conjugated to a detectable compound may be added to the well. Further, instead of coating the well with the antigen, the antibody may be coated to the well. In this case, a second antibody conjugated to a detectable compound may be added following the addition of the antigen of interest to the coated well. One of skill in the art would be knowledgeable as to the parameters that can be modified to increase the signal detected as well as other variations of ELISAs known in the art. For further discussion regarding ELISAs see, e.g., Ausubel et al, eds, (1994), Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, section 11.2.1.

The binding affinity of an antibody to an antigen and the off-rate of an antibody-antigen interaction can be determined by competitive binding assays. One example of a competitive binding assay is a radioimmunoassay comprising the incubation of labeled antigen (e.g., 3H or 125I) with the antibody of interest in the presence of increasing amounts of unlabeled antigen, and the detection of the antibody bound to the labeled antigen. The affinity of the antibody of interest for a particular antigen and the binding off-rates can be determined from the data by scatchard plot analysis. Competition with a second antibody can also be determined using radioimmunoassays. In this case, the antigen is incubated with antibody of interest conjugated to a labeled compound (e.g., 3H or 125I) in the presence of increasing amounts of an unlabeled second antibody.

Antibodies of the invention may be characterized using immunocytochemisty methods on cells (e.g., mammalian cells, such as CHO cells) transfected with a vector enabling the expression of an antigen or with vector alone using techniques commonly known in the art. Antibodies that bind antigen transfected cells, but not vector-only transfected cells, are antigen specific.

Therapeutic Uses

Tables 1D.1 and 1D.2 also provide information regarding biological activities and preferred therapeutic uses (i.e. see, “Preferred Indications” column) for polynucleotides and polypeptides of the invention (including antibodies, agonists, and/or antagonists thereof). Tables 1D.1 and 1D.2 also provide information regarding assays which may be used to test polynucleotides and polypeptides of the invention (including antibodies, agonists, and/or antagonists thereof) for the corresponding biological activities. The first column of Table 1D.1 (“Gene No.”) provides the gene number in the application for each clone identifier. The second column of Table 1D.1 (“cDNA Clone ID”) provides the unique clone identifier for each clone as previously described and indicated in Table 1A, Table 1B, and Table 1C. The third column of Table 1D.1 (“AA SEQ ID NO:Y”) indicates the Sequence Listing SEQ ID Number for polypeptide sequences encoded by the corresponding cDNA clones (also as indicated in Table 1A, Table 1B, and Table 2). The fourth column (“Biological Activity”) indicates a biological activity corresponding to the indicated polypeptides (or polynucleotides encoding said polypeptides). The first column of Table 1D.2 (“Biological Activity No.”) provides the biological activity number corresponding to each biological activity recited in the fourth column of Table 1D.1. The second column of Table 1D.2 (“Biological Activity”) indicates the biological activity corresponding to each biological activity number (also as indicated in Table 1D.1). The third column (“Exemplary Activity Assay”) further describes the corresponding biological activity and also provides information pertaining to the various types of assays which may be performed to test, demonstrate, or quantify the corresponding biological activity.

The present invention is further directed to antibody-based therapies which involve administering antibodies of the invention to an animal, preferably a mammal, and most preferably a human, patient for treating one or more of the disclosed diseases, disorders, or conditions. Therapeutic compounds of the invention include, but are not limited to, antibodies of the invention (including fragments, analogs and derivatives thereof as described herein) and nucleic acids encoding antibodies of the invention (including fragments, analogs and derivatives thereof and anti-idiotypic antibodies as described herein). The antibodies of the invention can be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate diseases, disorders or conditions associated with aberrant expression and/or activity of a polypeptide of the invention, including, but not limited to, immune diseases and disorders. The treatment and/or prevention of immune diseases and disorders associated with aberrant expression and/or activity of a polypeptide of the invention includes, but is not limited to, alleviating symptoms associated with immune diseases and disorders. Antibodies of the invention may be provided in pharmaceutically acceptable compositions as known in the art or as described herein.

In a specific and preferred embodiment, the present invention is directed to antibody-based therapies which involve administering antibodies of the invention to an animal, preferably a mammal, and most preferably a human, patient for treating immune diseases and disorders. Therapeutic compounds of the invention include, but are not limited to, antibodies of the invention (e.g., antibodies directed to the full length protein expressed on the cell surface of a mammalian cell; antibodies directed to an epitope of a polypeptide of the invention (such as, for example, a predicted linear epitope shown in Table 1B; or a conformational epitope, including fragments, analogs and derivatives thereof as described herein) and nucleic acids encoding antibodies of the invention (including fragments, analogs and derivatives thereof and anti-idiotypic antibodies as described herein). The antibodies of the invention can be used to detect, diagnose, prevent, treat, prognosticate, and/or ameliorate immune diseases, disorders or conditions associated with aberrant expression and/or activity of a polypeptide of the invention. The treatment and/or prevention of immune diseases, disorders, or conditions associated with aberrant expression and/or activity of a polypeptide of the invention includes, but is not limited to, alleviating symptoms associated with those diseases, disorders or conditions. Antibodies of the invention may be provided in pharmaceutically acceptable compositions as known in the art or as described herein.

A summary of the ways in which the antibodies of the present invention may be used therapeutically includes binding polynucleotides or polypeptides of the present invention locally or systemically in the body or by direct cytotoxicity of the antibody, e.g. as mediated by complement (CDC) or by effector cells (ADCC). Some of these approaches are described in more detail below. Armed with the teachings provided herein, one of ordinary skill in the art will know how to use the antibodies of the present invention for diagnostic, monitoring or therapeutic purposes without undue experimentation.

The antibodies of this invention may be advantageously utilized in combination with other monoclonal or chimeric antibodies, or with lymphokines or hematopoietic growth factors (such as, e.g., IL-2, IL-3 and IL-7), for example, which serve to increase the number or activity of effector cells which interact with the antibodies.

The antibodies of the invention may be administered alone or in combination with other types of treatments (e.g., radiation therapy, chemotherapy, hormonal therapy, immunotherapy and anti-tumor agents). Generally, administration of products of a species origin or species reactivity (in the case of antibodies) that is the same species as that of the patient is preferred. Thus, in a preferred embodiment, human antibodies, fragments derivatives, analogs, or nucleic acids, are administered to a human patient for therapy or prophylaxis.

It is preferred to use high affinity and/or potent in vivo inhibiting and/or neutralizing antibodies against polypeptides or polynucleotides of the present invention, fragments or regions thereof, for both immunoassays directed to and therapy of immune diseases and disorders related to polynucleotides or polypeptides, including fragments thereof, of the present invention. Such antibodies, fragments, or regions, will preferably have an affinity for polynucleotides or polypeptides of the invention, including fragments thereof. Preferred binding affinities include those with a dissociation constant or Kd less than 5×10⁻² M, 10⁻² M, 5×10⁻³ M, 10⁻³ M, 5×10⁻⁴ M, 10⁻⁴ M, 5×10⁻⁵ M, 10⁻⁵ M, 5×10⁻⁶ M, 10⁻⁶ M, 5×10⁻⁷ M, 10⁻⁷ M, 5×10⁻⁸ M, 10⁻⁸ M, 5×10⁻⁹ M, 10⁻⁹ M, 5×10⁻¹⁰ M ⁻¹⁰ M, 5×10⁻¹¹ M, 10⁻¹¹ M, 5×10⁻¹² M, 10⁻¹² M, 5×10⁻¹³ M, 10⁻¹³M 5×10⁻¹⁴ M, 10⁻¹⁴ M, 5×10⁻¹⁵ M, and 10⁻¹⁵ M.

Gene Therapy

In a specific embodiment, nucleic acids comprising sequences encoding antibodies or functional derivatives thereof, are administered to treat, inhibit or prevent a immune disease or disorder associated with aberrant expression and/or activity of a polypeptide of the invention, by way of gene therapy. Gene therapy refers to therapy performed by the administration to a subject of an expressed or expressible nucleic acid. In this embodiment of the invention, the nucleic acids produce their encoded protein that mediates a therapeutic effect.

Any of the methods for gene therapy available in the art can be used according to the present invention. Exemplary methods are described below.

For general reviews of the methods of gene therapy, see Goldspiel et al., Clinical Pharmacy 12:488-505 (1993); Wu and Wu, Biotherapy 3:87-95 (1991); Tolstoshev, Ann. Rev. Pharmacol. Toxicol. 32:573-596 (1993); Mulligan, Science 260:926-932 (1993); and Morgan and Anderson, Ann. Rev. Biochem. 62:191-217 (1993); May, TIBTECH 11(5):155-215 (1993). Methods commonly known in the art of recombinant DNA technology which can be used are described in Ausubel et al. (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993); and Kriegler, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990).

In a preferred embodiment, the compound comprises nucleic acid sequences encoding an antibody, said nucleic acid sequences being part of expression vectors that express the antibody or fragments or chimeric proteins or heavy or light chains thereof in a suitable host. In particular, such nucleic acid sequences have promoters operably linked to the antibody coding region, said promoter being inducible or constitutive, and, optionally, tissue-specific. In another particular embodiment, nucleic acid molecules are used in which the antibody coding sequences and any other desired sequences are flanked by regions that promote homologous recombination at a desired site in the genome, thus providing for intrachromosomal expression of the antibody encoding nucleic acids (Koller and Smithies, Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); Zijlstra et al., Nature 342:435-438 (1989). In specific embodiments, the expressed antibody molecule is a single chain antibody; alternatively, the nucleic acid sequences include sequences encoding both the heavy and light chains, or fragments thereof, of the antibody.

Delivery of the nucleic acids into a patient may be either direct, in which case the patient is directly exposed to the nucleic acid or nucleic acid-carrying vectors, or indirect, in which case, cells are first transformed with the nucleic acids in vitro, then transplanted into the patient. These two approaches are known, respectively, as in vivo or ex vivo gene therapy.

In a specific embodiment, the nucleic acid sequences are directly administered in vivo, where it is expressed to produce the encoded product. This can be accomplished by any of numerous methods known in the art, e.g., by constructing them as part of an appropriate nucleic acid expression vector and administering it so that they become intracellular, e.g., by infection using defective or attenuated retrovirals or other viral vectors (see U.S. Pat. No. 4,980,286), or by direct injection of naked DNA, or by use of microparticle bombardment (e.g., a gene gun; Biolistic, DUPONT™), or coating with lipids or cell-surface receptors or transfecting agents, encapsulation in liposomes, microparticles, or microcapsules, or by administering them in linkage to a peptide which is known to enter the nucleus, by administering it in linkage to a ligand subject to receptor-mediated endocytosis (see, e.g., Wu and Wu, J. Biol. Chem. 262:4429-4432 (1987)) (which can be used to target cell types specifically expressing the receptors), etc. In another embodiment, nucleic acid-ligand complexes can be formed in which the ligand comprises a fusogenic viral peptide to disrupt endosomes, allowing the nucleic acid to avoid lysosomal degradation. In yet another embodiment, the nucleic acid can be targeted in vivo for cell specific uptake and expression, by targeting a specific receptor (see, e.g., PCT Publications WO 92/06180; WO 92/22635; WO92/20316; WO93/14188, WO 93/20221). Alternatively, the nucleic acid can be introduced intracellularly and incorporated within host cell DNA for expression, by homologous recombination (Koller and Smithies, Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); Zijlstra et al., Nature 342:435-438 (1989)).

In a specific embodiment, viral vectors which contain nucleic acid sequences encoding an antibody of the invention are used. For example, a retroviral vector can be used (see Miller et al., Meth. Enzymol. 217:581-599 (1993)). These retroviral vectors contain the components necessary for the correct packaging of the viral genome and integration into the host cell DNA. The nucleic acid sequences encoding the antibody to be used in gene therapy are cloned into one or more vectors, which facilitates delivery of the gene into a patient. More detail about retroviral vectors can be found in Boesen et al., Biotherapy 6:291-302 (1994), which describes the use of a retroviral vector to deliver the mdr1 gene to hematopoietic stem cells in order to make the stem cells more resistant to chemotherapy. Other references illustrating the use of retroviral vectors in gene therapy are: Clowes et al., J. Clin. Invest. 93:644-651 (1994); Kiem et al., Blood 83:1467-1473 (1994); Salmons and Gunzberg, Human Gene Therapy 4:129-141 (1993); and Grossman and Wilson, Curr. Opin. in Genetics and Devel. 3:110-114 (1993).

Adenoviruses are other viral vectors that can be used in gene therapy. Adenoviruses are especially attractive vehicles for delivering genes to respiratory epithelia. Adenoviruses naturally infect respiratory epithelia where they cause a mild disease. Other targets for adenovirus-based delivery systems are liver, the central nervous system, endothelial cells, and muscle. Adenoviruses have the advantage of being capable of infecting non-dividing cells. Kozarsky and Wilson, Current Opinion in Genetics and Development 3:499-503 (1993) present a review of adenovirus-based gene therapy. Bout et al., Human Gene Therapy 5:3-10 (1994) demonstrated the use of adenovirus vectors to transfer genes to the respiratory epithelia of rhesus monkeys. Other instances of the use of adenoviruses in gene therapy can be found in Rosenfeld et al., Science 252:431-434 (1991); Rosenfeld et al., Cell 68:143-155 (1992); Mastrangeli et al., J. Clin. Invest. 91:225-234 (1993); PCT Publication WO94/12649; and Wang, et al., Gene Therapy 2:775-783 (1995). In a preferred embodiment, adenovirus vectors are used.

Adeno-associated virus (AAV) has also been proposed for use in gene therapy (Walsh et al., Proc. Soc. Exp. Biol. Med. 204:289-300 (1993); U.S. Pat. No. 5,436,146).

Another approach to gene therapy involves transferring a gene to cells in tissue culture by such methods as electroporation, lipofection, calcium phosphate mediated transfection, or viral infection. Usually, the method of transfer includes the transfer of a selectable marker to the cells. The cells are then placed under selection to isolate those cells that have taken up and are expressing the transferred gene. Those cells are then delivered to a patient.

In this embodiment, the nucleic acid is introduced into a cell prior to administration in vivo of the resulting recombinant cell. Such introduction can be carried out by any method known in the art, including but not limited to transfection, electroporation, microinjection, infection with a viral or bacteriophage vector containing the nucleic acid sequences, cell fusion, chromosome-mediated gene transfer, microcell-mediated gene transfer, spheroplast fusion, etc. Numerous techniques are known in the art for the introduction of foreign genes into cells (see, e.g., Loeffler and Behr, Meth. Enzymol. 217:599-618 (1993); Cohen et al., Meth. Enzymol. 217:618-644 (1993); Cline, Pharmac. Ther. 29:69-92m (1985) and may be used in accordance with the present invention, provided that the necessary developmental and physiological functions of the recipient cells are not disrupted. The technique should provide for the stable transfer of the nucleic acid to the cell, so that the nucleic acid is expressible by the cell and preferably heritable and expressible by its cell progeny.

The resulting recombinant cells can be delivered to a patient by various methods known in the art. Recombinant blood cells (e.g., hematopoietic stem or progenitor cells) are preferably administered intravenously. The amount of cells envisioned for use depends on the desired effect, patient state, etc., and can be determined by one skilled in the art.

Cells into which a nucleic acid can be introduced for purposes of gene therapy encompass any desired, available cell type, and include but are not limited to epithelial cells, endothelial cells, keratinocytes, fibroblasts, muscle cells, hepatocytes; blood cells such as T lymphocytes, B lymphocytes, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes, granulocytes; various stem or progenitor cells, in particular hematopoietic stem or progenitor cells, e.g., as obtained from bone marrow, umbilical cord blood, peripheral blood, fetal liver, etc.

In a preferred embodiment, the cell used for gene therapy is autologous to the patient.

In an embodiment in which recombinant cells are used in gene therapy, nucleic acid sequences encoding an antibody are introduced into the cells such that they are expressible by the cells or their progeny, and the recombinant cells are then administered in vivo for therapeutic effect. In a specific embodiment, stem or progenitor cells are used. Any stem and/or progenitor cells which can be isolated and maintained in vitro can potentially be used in accordance with this embodiment of the present invention (see e.g. PCT Publication WO 94/08598; Stemple and Anderson, Cell 71:973-985 (1992); Rheinwald, Meth. Cell Bio. 21A:229 (1980); and Pittelkow and Scott, Mayo Clinic Proc. 61:771 (1986)).

In a specific embodiment, the nucleic acid to be introduced for purposes of gene therapy comprises an inducible promoter operably linked to the coding region, such that expression of the nucleic acid is controllable by the presence or absence of an appropriate inducer of transcription.

Demonstration of Therapeutic or Prophylactic Activity

The compounds or pharmaceutical compositions of the invention are preferably tested in vitro, and then in vivo for the desired therapeutic or prophylactic activity, prior to use in humans. For example, in vitro assays to demonstrate the therapeutic or prophylactic utility of a compound or pharmaceutical composition include, the effect of a compound on a cell line or a patient tissue sample. The effect of the compound or composition on the cell line and/or tissue sample can be determined utilizing techniques known to those of skill in the art including, but not limited to, rosette formation assays and cell lysis assays. In accordance with the invention, in vitro assays which can be used to determine whether administration of a specific compound is indicated, include in vitro cell culture assays in which a patient tissue sample is grown in culture, and exposed to or otherwise administered a compound, and the effect of such compound upon the tissue sample is observed.

Therapeutic/Prophylactic Administration and Composition

The invention provides methods of treatment, inhibition and prophylaxis by administration to a subject of an effective amount of a compound or pharmaceutical composition of the invention, preferably a polypeptide or antibody of the invention. In a preferred embodiment, the compound is substantially purified (e.g., substantially free from substances that limit its effect or produce undesired side-effects). The subject is preferably an animal, including but not limited to animals such as cows, pigs, horses, chickens, cats, dogs, etc., and is preferably a mammal, and most preferably human.

Formulations and methods of administration that can be employed when the compound comprises a nucleic acid or an immunoglobulin are described above; additional appropriate formulations and routes of administration can be selected from among those described herein below.

Various delivery systems are known and can be used to administer a compound of the invention, e.g., encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the compound, receptor-mediated endocytosis (see, e.g., Wu and Wu, J. Biol. Chem. 262:4429-4432 (1987)), construction of a nucleic acid as part of a retroviral or other vector, etc. Methods of introduction include but are not limited to intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The compounds or compositions may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.) and may be administered together with other biologically active agents. Administration can be systemic or local. In addition, it may be desirable to introduce the pharmaceutical compounds or compositions of the invention into the central nervous system by any suitable route, including intraventricular and intrathecal injection; intraventricular injection may be facilitated by an intraventricular catheter, for example, attached to a reservoir, such as an Ommaya reservoir. Pulmonary administration can also be employed, e.g., by use of an inhaler or nebulizer, and formulation with an aerosolizing agent.

In a specific embodiment, it may be desirable to administer the pharmaceutical compounds or compositions of the invention locally to the area in need of treatment; this may be achieved by, for example, and not by way of limitation, local infusion during surgery, topical application, e.g., in conjunction with a wound dressing after surgery, by injection, by means of a catheter, by means of a suppository, or by means of an implant, said implant being of a porous, non-porous, or gelatinous material, including membranes, such as sialastic membranes, or fibers. Preferably, when administering a protein, including an antibody, of the invention, care must be taken to use materials to which the protein does not absorb.

In another embodiment, the compound or composition can be delivered in a vesicle, in particular a liposome (see Langer, Science 249:1527-1533 (1990); Treat et al., in Liposomes in the Therapy of Infectious Disease and Cancer, Lopez-Berestein and Fidler (eds.), Liss, New York, pp. 353-365 (1989); Lopez-Berestein, ibid., pp. 317-327; see generally ibid.)

In yet another embodiment, the compound or composition can be delivered in a controlled release system. In one embodiment, a pump may be used (see Langer, supra; Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987); Buchwald et al., Surgery 88:507 (1980); Saudek et al., N. Engl. J. Med. 321:574 (1989)). In another embodiment, polymeric materials can be used (see Medical Applications of Controlled Release, Langer and Wise (eds.), CRC Pres., Boca Raton, Fla. (1974); Controlled Drug Bioavailability, Drug Product Design and Performance, Smolen and Ball (eds.), Wiley, New York (1984); Ranger and Peppas, J., Macromol. Sci. Rev. Macromol. Chem. 23:61 (1983); see also Levy et al., Science 228:190 (1985); During et al., Ann. Neurol. 25:351 (1989); Howard et al., J. Neurosurg. 71:105 (1989)). In yet another embodiment, a controlled release system can be placed in proximity of the therapeutic target, e.g., the brain, thus requiring only a fraction of the systemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138 (1984)).

Other controlled release systems are discussed in the review by Langer (Science 249:1527-1533 (1990)).

In a specific embodiment where the compound of the invention is a nucleic acid encoding a protein, the nucleic acid can be administered in vivo to promote expression of its encoded protein, by constructing it as part of an appropriate nucleic acid expression vector and administering it so that it becomes intracellular, e.g., by use of a retroviral vector (see U.S. Pat. No. 4,980,286), or by direct injection, or by use of microparticle bombardment (e.g., a gene gun; Biolistic, DUPONT™), or coating with lipids or cell-surface receptors or transfecting agents, or by administering it in linkage to a homeobox-like peptide which is known to enter the nucleus (see e.g., Joliot et al., Proc. Natl. Acad. Sci. USA 88:1864-1868 (1991)), etc. Alternatively, a nucleic acid can be introduced intracellularly and incorporated within host cell DNA for expression, by homologous recombination.

The present invention also provides pharmaceutical compositions. Such compositions comprise a therapeutically effective amount of a compound, and a pharmaceutically acceptable carrier. In a specific embodiment, the term “pharmaceutically acceptable” means approved by a regulatory agency of the Federal or a state government or listed in the U.S. Pharmacopeia or other generally recognized pharmacopeia for use in animals, and more particularly in humans. The term “carrier” refers to a diluent, adjuvant, excipient, or vehicle with which the therapeutic is administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water is a preferred carrier when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be employed as liquid carriers, particularly for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol and the like. The composition, if desired, can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. These compositions can take the form of solutions, suspensions, emulsion, tablets, pills, capsules, powders, sustained-release formulations and the like. The composition can be formulated as a suppository, with traditional binders and carriers such as triglycerides. Oral formulation can include standard carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate, etc. Examples of suitable pharmaceutical carriers are described in “Remington's Pharmaceutical Sciences” by E. W. Martin. Such compositions will contain a therapeutically effective amount of the compound, preferably in purified form, together with a suitable amount of carrier so as to provide the form for proper administration to the patient. The formulation should suit the mode of administration.

In a preferred embodiment, the composition is formulated in accordance with routine procedures as a pharmaceutical composition adapted for intravenous administration to human beings. Typically, compositions for intravenous administration are solutions in sterile isotonic aqueous buffer. Where necessary, the composition may also include a solubilizing agent and a local anesthetic such as lignocaine to ease pain at the site of the injection. Generally, the ingredients are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilized powder or water free concentrate in a hermetically sealed container such as an ampoule or sachette indicating the quantity of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline. Where the composition is administered by injection, an ampoule of sterile water for injection or saline can be provided so that the ingredients may be mixed prior to administration.

The compounds of the invention can be formulated as neutral or salt forms. Pharmaceutically acceptable salts include those formed with anions such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with cations such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, procaine, etc.

The amount of the compound of the invention which will be effective in the treatment, inhibition and prevention of a disease or disorder associated with aberrant expression and/or activity of a polypeptide of the invention can be determined by standard clinical techniques. In addition, in vitro assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed in the formulation will also depend on the route of administration, and the seriousness of the disease or disorder, and should be decided according to the judgment of the practitioner and each patient's circumstances. Effective doses may be extrapolated from dose-response curves derived from in vitro or animal model test systems.

For antibodies, the dosage administered to a patient is typically 0.1 mg/kg to 100 mg/kg of the patient's body weight. Preferably, the dosage administered to a patient is between 0.1 mg/kg and 20 mg/kg of the patient's body weight, more preferably 1 mg/kg to 10 mg/kg of the patient's body weight. Generally, human antibodies have a longer half-life within the human body than antibodies from other species due to the immune response to the foreign polypeptides. Thus, lower dosages of human antibodies and less frequent administration is often possible. Further, the dosage and frequency of administration of antibodies of the invention may be reduced by enhancing uptake and tissue penetration (e.g., into the brain) of the antibodies by modifications such as, for example, lipidation.

The invention also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the pharmaceutical compositions of the invention. Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration.

Diagnosis and Imaging

Labeled antibodies, and derivatives and analogs thereof, which specifically bind to a polypeptide of interest can be used for diagnostic purposes to detect, diagnose, prognosticate, or monitor immune diseases, disorders, and/or conditions associated with the aberrant expression and/or activity of a polypeptide of the invention. The invention provides for the detection of aberrant expression of a polypeptide of interest, comprising (a) assaying the expression of the polypeptide of interest in cells or body fluid of an individual using one or more antibodies specific to the polypeptide interest and (b) comparing the level of gene expression with a standard gene expression level, whereby an increase or decrease in the assayed polypeptide gene expression level compared to the standard expression level is indicative of aberrant expression.

The invention provides a diagnostic assay for diagnosing an immune disease or disorder, comprising (a) assaying the expression of the polypeptide of interest in cells or body fluid of an individual using one or more antibodies specific to the polypeptide interest and (b) comparing the level of gene expression with a standard gene expression level, whereby an increase or decrease in the assayed polypeptide gene expression level compared to the standard expression level is indicative of a particular immune disease or disorder. With respect to immunogenic cancers, the presence of a relatively high amount of transcript in biopsied tissue from an individual may indicate a predisposition for the development of the disease, or may provide a means for detecting the disease prior to the appearance of actual clinical symptoms. A more definitive diagnosis of this type may allow health professionals to employ preventative measures or aggressive treatment earlier thereby preventing the development or further progression of the immunogenic cancer.

Antibodies of the invention can be used to assay protein levels in a biological sample using classical immunohistological methods known to those of skill in the art (e.g., see Jalkanen et al., J. Cell. Biol. 101:976-985 (1985); Jalkanen et al., J. Cell. Biol. 105:3087-3096 (1987)). Other antibody-based methods useful for detecting protein gene expression include immunoassays, such as the enzyme linked immunosorbent assay (ELISA) and the radioimmunoassay (RIA). Suitable antibody assay labels are known in the art and include enzyme labels, such as, glucose oxidase; radioisotopes, such as iodine (125I, 121I), carbon (14C), sulfur (35S), tritium (3H), indium (112In), and technetium (99Tc); luminescent labels, such as luminol; and fluorescent labels, such as fluorescein and rhodamine, and biotin.

One facet of the invention is the detection and diagnosis of a disease or disorder associated with aberrant expression of a polypeptide of interest in an animal, preferably a mammal and most preferably a human. In one embodiment, diagnosis comprises: a) administering (for example, parenterally, subcutaneously, or intraperitoneally) to a subject an effective amount of a labeled molecule which specifically binds to the polypeptide of interest; b) waiting for a time interval following the administering for permitting the labeled molecule to preferentially concentrate at sites in the subject where the polypeptide is expressed (and for unbound labeled molecule to be cleared to background level); c) determining background level; and d) detecting the labeled molecule in the subject, such that detection of labeled molecule above the background level indicates that the subject has a particular disease or disorder associated with aberrant expression of the polypeptide of interest. Background level can be determined by various methods including, comparing the amount of labeled molecule detected to a standard value previously determined for a particular system.

It will be understood in the art that the size of the subject and the imaging system used will determine the quantity of imaging moiety needed to produce diagnostic images. In the case of a radioisotope moiety, for a human subject, the quantity of radioactivity injected will normally range from about 5 to 20 millicuries of 99 mTc. The labeled antibody or antibody fragment will then preferentially accumulate at the location of cells which contain the specific protein. In vivo tumor imaging is described in S. W. Burchiel et al, “Immunopharmacokinetics of Radiolabeled Antibodies and Their Fragments.” (Chapter 13 in Tumor Imaging: The Radiochemical Detection of Cancer, S. W. Burchiel and B. A. Rhodes, eds., Masson Publishing Inc. (1982)).

Depending on several variables, including the type of label used and the mode of administration, the time interval following the administration for permitting the labeled molecule to preferentially concentrate at sites in the subject and for unbound labeled molecule to be cleared to background level is 6 to 48 hours or 6 to 24 hours or 6 to 12 hours. In another embodiment the time interval following administration is 5 to 20 days or 5 to 10 days.

In an embodiment, monitoring of the disease or disorder is carried out by repeating the method for diagnosing the disease or disease, for example, one month after initial diagnosis, six months after initial diagnosis, one year after initial diagnosis, etc.

Presence of the labeled molecule can be detected in the patient using methods known in the art for in vivo scanning. These methods depend upon the type of label used. Skilled artisans will be able to determine the appropriate method for detecting a particular label. Methods and devices that may be used in the diagnostic methods of the invention include, but are not limited to, computed tomography (CT), whole body scan such as position emission tomography (PET), magnetic resonance imaging (MRI), and sonography.

In a specific embodiment, the molecule is labeled with a radioisotope and is detected in the patient using a radiation responsive surgical instrument (Thurston et al., U.S. Pat. No. 5,441,050). In another embodiment, the molecule is labeled with a fluorescent compound and is detected in the patient using a fluorescence responsive scanning instrument. In another embodiment, the molecule is labeled with a positron emitting metal and is detected in the patent using positron emission-tomography. In yet another embodiment, the molecule is labeled with a paramagnetic label and is detected in a patient using magnetic resonance imaging (MRI).

Kits

The present invention provides kits that can be used in the above methods. In one embodiment, a kit comprises an antibody of the invention, preferably a purified antibody, in one or more containers. In a specific embodiment, the kits of the present invention contain a substantially isolated polypeptide comprising an epitope that is specifically immunoreactive with an antibody included in the kit. Preferably, the kits of the present invention further comprise a control antibody that does not react with the polypeptide of interest. In another specific embodiment, the kits of the present invention contain a means for detecting the binding of an antibody to a polypeptide of interest (e.g., the antibody may be conjugated to a detectable substrate such as a fluorescent compound, an enzymatic substrate, a radioactive compound or a luminescent compound, or a second antibody which recognizes the first antibody may be conjugated to a detectable substrate).

In another specific embodiment of the present invention, the kit is a diagnostic kit for use in screening serum containing antibodies specific against proliferative and/or cancerous polynucleotides and polypeptides. Such a kit may include a control antibody that does not react with the polypeptide of interest. Such a kit may include a substantially isolated polypeptide antigen comprising an epitope that is specifically immunoreactive with at least one anti-polypeptide antigen antibody. Further, such a kit includes means for detecting the binding of said antibody to the antigen (e.g., the antibody may be conjugated to a fluorescent compound such as fluorescein or rhodamine which can be detected by flow cytometry). In specific embodiments, the kit may include a recombinantly produced or chemically synthesized polypeptide antigen. The polypeptide antigen of the kit may also be attached to a solid support.

In a more specific embodiment the detecting means of the above-described kit includes a solid support to which said polypeptide antigen is attached. Such a kit may also include a non-attached reporter-labeled anti-human antibody. In this embodiment, binding of the antibody to the polypeptide antigen can be detected by binding of the said reporter-labeled antibody.

In an additional embodiment, the invention includes a diagnostic kit for use in screening serum containing antigens of the polypeptide of the invention. The diagnostic kit includes a substantially isolated antibody specifically immunoreactive with polypeptide or polynucleotide antigens, and means for detecting the binding of the polynucleotide or polypeptide antigen to the antibody. In one embodiment, the antibody is attached to a solid support. In a specific embodiment, the antibody may be a monoclonal antibody. The detecting means of the kit may include a second, labeled monoclonal antibody. Alternatively, or in addition, the detecting means may include a labeled, competing antigen.

In one diagnostic configuration, test serum is reacted with a solid phase reagent having a surface-bound antigen obtained by the methods of the present invention. After binding with specific antigen antibody to the reagent and removing unbound serum components by washing, the reagent is reacted with reporter-labeled anti-human antibody to bind reporter to the reagent in proportion to the amount of bound anti-antigen antibody on the solid support. The reagent is again washed to remove unbound labeled antibody, and the amount of reporter associated with the reagent is determined. Typically, the reporter is an enzyme which is detected by incubating the solid phase in the presence of a suitable fluorometric, luminescent or colorimetric substrate (Sigma, St. Louis, Mo.).

The solid surface reagent in the above assay is prepared by known techniques for attaching protein material to solid support material, such as polymeric beads, dip sticks, 96-well plate or filter material. These attachment methods generally include non-specific adsorption of the protein to the support or covalent attachment of the protein, typically through a free amine group, to a chemically reactive group on the solid support, such as an activated carboxyl, hydroxyl, or aldehyde group. Alternatively, streptavidin coated plates can be used in conjunction with biotinylated antigen(s).

Thus, the invention provides an assay system or kit for carrying out this diagnostic method. The kit generally includes a support with surface-bound recombinant antigens, and a reporter-labeled anti-human antibody for detecting surface-bound anti-antigen antibody.

Uses of the Polynucleotides

Each of the polynucleotides identified herein can be used in numerous ways as reagents. The following description should be considered exemplary and utilizes known techniques.

The polynucleotides of the present invention are useful for chromosome identification. There exists an ongoing need to identify new chromosome markers, since few chromosome marking reagents, based on actual sequence data (repeat polymorphisms), are presently available. Each sequence is specifically targeted to and can hybridize with a particular location on an individual human chromosome, thus each polynucleotide of the present invention can routinely be used as a chromosome marker using techniques known in the art. Table 1B.1, column 8 provides the chromosome location of some of the polynucleotides of the invention.

Briefly, sequences can be mapped to chromosomes by preparing PCR primers (preferably at least 15 by (e.g., 15-25 bp) from the sequences shown in SEQ ID NO:X. Primers can optionally be selected using computer analysis so that primers do not span more than one predicted exon in the genomic DNA. These primers are then used for PCR screening of somatic cell hybrids containing individual human chromosomes. Only those hybrids containing the human gene corresponding to SEQ ID NO:X will yield an amplified fragment.

Similarly, somatic hybrids provide a rapid method of PCR mapping the polynucleotides to particular chromosomes. Three or more clones can be assigned per day using a single thermal cycler. Moreover, sublocalization of the polynucleotides can be achieved with panels of specific chromosome fragments. Other gene mapping strategies that can be used include in situ hybridization, prescreening with labeled flow-sorted chromosomes, preselection by hybridization to construct chromosome specific-cDNA libraries, and computer mapping techniques (See, e.g., Shuler, Trends Biotechnol 16:456-459 (1998) which is hereby incorporated by reference in its entirety).

Precise chromosomal location of the polynucleotides can also be achieved using fluorescence in situ hybridization (FISH) of a metaphase chromosomal spread. This technique uses polynucleotides as short as 500 or 600 bases; however, polynucleotides 2,000-4,000 by are preferred. For a review of this technique, see Verma et al., “Human Chromosomes: a Manual of Basic Techniques,” Pergamon Press, New York (1988).

For chromosome mapping, the polynucleotides can be used individually (to mark a single chromosome or a single site on that chromosome) or in panels (for marking multiple sites and/or multiple chromosomes).

Thus, the present invention also provides a method for chromosomal localization which involves (a) preparing PCR primers from the polynucleotide sequences in Table 1B and/or Table 2 and SEQ ID NO:X and (b) screening somatic cell hybrids containing individual chromosomes.

The polynucleotides of the present invention would likewise be useful for radiation hybrid mapping, HAPPY mapping, and long range restriction mapping. For a review of these techniques and others known in the art, see, e.g. Dear, “Genome Mapping: A Practical Approach,” IRL Press at Oxford University Press, London (1997); Aydin, J. Mol. Med. 77:691-694 (1999); Hacia et al., Mol. Psychiatry. 3:483-492 (1998); Herrick et al., Chromosome Res. 7:409-423 (1999); Hamilton et al., Methods Cell Biol. 62:265-280 (2000); and/or Ott, J. Hered. 90:68-70 (1999) each of which is hereby incorporated by reference in its entirety.

Once a polynucleotide has been mapped to a precise chromosomal location, the physical position of the polynucleotide can be used in linkage analysis. Linkage analysis establishes coinheritance between a chromosomal location and presentation of a particular disease. (Disease mapping data are found, for example, in V. McKusick, Mendelian Inheritance in Man (available on line through Johns Hopkins University Welch Medical Library)). Column 9 of Table 1B.1 provides an OMIM reference identification number of diseases associated with the cytologic band disclosed in column 8 of Table 1B.1, as determined using techniques described herein and by reference to Table 5. Assuming 1 megabase mapping resolution and one gene per 20 kb, a cDNA precisely localized to a chromosomal region associated with the disease could be one of 50-500 potential causative genes.

Thus, once coinheritance is established, differences in a polynucleotide of the invention and the corresponding gene between affected and unaffected individuals can be examined. First, visible structural alterations in the chromosomes, such as deletions or translocations, are examined in chromosome spreads or by PCR. If no structural alterations exist, the presence of point mutations are ascertained. Mutations observed in some or all affected individuals, but not in normal individuals, indicates that the mutation may cause the disease. However, complete sequencing of the polypeptide and the corresponding gene from several normal individuals is required to distinguish the mutation from a polymorphism. If a new polymorphism is identified, this polymorphic polypeptide can be used for further linkage analysis.

Furthermore, increased or decreased expression of the gene in affected individuals as compared to unaffected individuals can be assessed using the polynucleotides of the invention. Any of these alterations (altered expression, chromosomal rearrangement, or mutation) can be used as a diagnostic or prognostic marker. Diagnostic and prognostic methods, kits and reagents encompassed by the present invention are briefly described below and more thoroughly elsewhere herein (see e.g., the sections labeled “Antibodies”, “Diagnostic Assays”, and “Methods for Detecting Diseases”).

Thus, the invention also provides a diagnostic method useful during diagnosis of a disorder, involving measuring the expression level of polynucleotides of the present invention in cells or body fluid from an individual and comparing the measured gene expression level with a standard level of polynucleotide expression level, whereby an increase or decrease in the gene expression level compared to the standard is indicative of a disorder. Additional non-limiting examples of diagnostic methods encompassed by the present invention are more thoroughly described elsewhere herein (see, e.g., Example 12).

In still another embodiment, the invention includes a kit for analyzing samples for the presence of proliferative and/or cancerous polynucleotides derived from a test subject. In a general embodiment, the kit includes at least one polynucleotide probe containing a nucleotide sequence that will specifically hybridize with a polynucleotide of the invention and a suitable container. In a specific embodiment, the kit includes two polynucleotide probes defining an internal region of the polynucleotide of the invention, where each probe has one strand containing a 31′ mer-end internal to the region. In a further embodiment, the probes may be useful as primers for polymerase chain reaction amplification.

Where a diagnosis of a related disorder, including, for example, diagnosis of a tumor, has already been made according to conventional methods, the present invention is useful as a prognostic indicator, whereby patients exhibiting enhanced or depressed polynucleotide of the invention expression will experience a worse clinical outcome relative to patients expressing the gene at a level nearer the standard level.

By “measuring the expression level of polynucleotides of the invention” is intended qualitatively or quantitatively measuring or estimating the level of the polypeptide of the invention or the level of the mRNA encoding the polypeptide of the invention in a first biological sample either directly (e.g., by determining or estimating absolute protein level or mRNA level) or relatively (e.g., by comparing to the polypeptide level or mRNA level in a second biological sample). Preferably, the polypeptide level or mRNA level in the first biological sample is measured or estimated and compared to a standard polypeptide level or mRNA level, the standard being taken from a second biological sample obtained from an individual not having the related disorder or being determined by averaging levels from a population of individuals not having a related disorder. As will be appreciated in the art, once a standard polypeptide level or mRNA level is known, it can be used repeatedly as a standard for comparison.

By “biological sample” is intended any biological sample obtained from an individual, body fluid, cell line, tissue culture, or other source that contains polypeptide of the present invention or the corresponding mRNA. As indicated, biological samples include body fluids (such as semen, lymph, vaginal pool, sera, plasma, urine, synovial fluid and spinal fluid) which contain the polypeptide of the present invention, and tissue sources found to express the polypeptide of the present invention. Methods for obtaining tissue biopsies and body fluids from mammals are well known in the art. Where the biological sample is to include mRNA, a tissue biopsy is the preferred source.

The method(s) provided above may preferably be applied in a diagnostic method and/or kits in which polynucleotides and/or polypeptides of the invention are attached to a solid support. In one exemplary method, the support may be a “gene chip” or a “biological chip” as described in U.S. Pat. Nos. 5,837,832, 5,874,219, and 5,856,174. Further, such a gene chip with polynucleotides of the invention attached may be used to identify polymorphisms between the isolated polynucleotide sequences of the invention, with polynucleotides isolated from a test subject. The knowledge of such polymorphisms (i.e. their location, as well as, their existence) would be beneficial in identifying disease loci for many disorders, such as for example, in neural disorders, immune system disorders, muscular disorders, reproductive disorders, gastrointestinal disorders, pulmonary disorders, digestive disorders, metabolic disorders, cardiovascular disorders, renal disorders, proliferative disorders, and/or cancerous diseases and conditions. Such a method is described in U.S. Pat. Nos. 5,858,659 and 5,856,104. The US Patents referenced supra are hereby incorporated by reference in their entirety herein.

The present invention encompasses polynucleotides of the present invention that are chemically synthesized, or reproduced as peptide nucleic acids (PNA), or according to other methods known in the art. The use of PNAs would serve as the preferred form if the polynucleotides of the invention are incorporated onto a solid support, or gene chip. For the purposes of the present invention, a peptide nucleic acid (PNA) is a polyamide type of DNA analog and the monomeric units for adenine, guanine, thymine and cytosine are available commercially (Perceptive Biosystems). Certain components of DNA, such as phosphorus, phosphorus oxides, or deoxyribose derivatives, are not present in PNAs. As disclosed by Nielsen et al., Science 254, 1497 (1991); and Egholm et al., Nature 365, 666 (1993), PNAs bind specifically and tightly to complementary DNA strands and are not degraded by nucleases. In fact, PNA binds more strongly to DNA than DNA itself does. This is probably because there is no electrostatic repulsion between the two strands, and also the polyamide backbone is more flexible. Because of this, PNA/DNA duplexes bind under a wider range of stringency conditions than DNA/DNA duplexes, making it easier to perform multiplex hybridization. Smaller probes can be used than with DNA due to the strong binding. In addition, it is more likely that single base mismatches can be determined with PNA/DNA hybridization because a single mismatch in a PNA/DNA 15-mer lowers the melting point (T_(m)) by 8°-20° C., vs. 4°-16° C. for the DNA/DNA 15-mer duplex. Also, the absence of charge groups in PNA means that hybridization can be done at low ionic strengths and reduce possible interference by salt during the analysis.

The compounds of the present invention have uses which include, but are not limited to, detecting cancer in mammals. In particular the invention is useful during diagnosis of pathological cell proliferative neoplasias which include, but are not limited to: acute myelogenous leukemias including acute monocytic leukemia, acute myeloblastic leukemia, acute promyelocytic leukemia, acute myelomonocytic leukemia, acute erythroleukemia, acute megakaryocytic leukemia, and acute undifferentiated leukemia, etc.; and chronic myelogenous leukemias including chronic myelomonocytic leukemia, chronic granulocytic leukemia, etc. Preferred mammals include monkeys, apes, cats, dogs, cows, pigs, horses, rabbits and humans. Particularly preferred are humans.

Pathological cell proliferative disorders are often associated with inappropriate activation of proto-oncogenes. (Gelmann, E. P. et al., “The Etiology of Acute Leukemia: Molecular Genetics and Viral Oncology,” in Neoplastic Diseases of the Blood, Vol 1., Wiernik, P. H. et al. eds., 161-182 (1985)). Neoplasias are now believed to result from the qualitative alteration of a normal cellular gene product, or from the quantitative modification of gene expression by insertion into the chromosome of a viral sequence, by chromosomal translocation of a gene to a more actively transcribed region, or by some other mechanism. (Gelmann et al., supra) It is likely that mutated or altered expression of specific genes is involved in the pathogenesis of some leukemias, among other tissues and cell types. (Gelmann et al., supra) Indeed, the human counterparts of the oncogenes involved in some animal neoplasias have been amplified or translocated in some cases of human leukemia and carcinoma. (Gelmann et al., supra)

For example, c-myc expression is highly amplified in the non-lymphocytic leukemia cell line HL-60. When HL-60 cells are chemically induced to stop proliferation, the level of c-myc is found to be down-regulated. (International Publication Number WO 91/15580). However, it has been shown that exposure of HL-60 cells to a DNA construct that is complementary to the 5′ end of c-myc or c-myb blocks translation of the corresponding mRNAs which downregulates expression of the c-myc or c-myb proteins and causes arrest of cell proliferation and differentiation of the treated cells. (International Publication Number WO 91/15580; Wickstrom et al., Proc. Natl. Acad. Sci. 85:1028 (1988); Anfossi et al., Proc. Natl. Acad. Sci. 86:3379 (1989)). However, the skilled artisan would appreciate the present invention's usefulness is not be limited to treatment, prevention, and/or prognosis of proliferative disorders of cells and tissues of hematopoietic origin, in light of the numerous cells and cell types of varying origins which are known to exhibit proliferative phenotypes.

In addition to the foregoing, a polynucleotide of the present invention can be used to control gene expression through triple helix formation or through antisense DNA or RNA. Antisense techniques are discussed, for example, in Okano, J. Neurochem. 56: 560 (1991); “Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988). Triple helix formation is discussed in, for instance Lee et al., Nucleic Acids Research 6: 3073 (1979); Cooney et al., Science 241: 456 (1988); and Dervan et al., Science 251: 1360 (1991). Both methods rely on binding of the polynucleotide to a complementary DNA or RNA. For these techniques, preferred polynucleotides are usually oligonucleotides 20 to 40 bases in length and complementary to either the region of the gene involved in transcription (triple helix—see Lee et al., Nucl. Acids Res. 6:3073 (1979); Cooney et al., Science 241:456 (1988); and Dervan et al., Science 251:1360 (1991)) or to the mRNA itself (antisense—Okano, J. Neurochem. 56:560 (1991); Oligodeoxy-nucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988)). Triple helix formation optimally results in a shut-off of RNA transcription from DNA, while antisense RNA hybridization blocks translation of an mRNA molecule into polypeptide. The oligonucleotide described above can also be delivered to cells such that the antisense RNA or DNA may be expressed in vivo to inhibit production of polypeptide of the present invention antigens. Both techniques are effective in model systems, and the information disclosed herein can be used to design antisense or triple helix polynucleotides in an effort to treat disease, and in particular, for the treatment of proliferative diseases and/or conditions. Non-limiting antisense and triple helix methods encompassed by the present invention are more thoroughly described elsewhere herein (see, e.g., the section labeled “Antisense and Ribozyme (Antagonists)”).

Polynucleotides of the present invention are also useful in gene therapy. One goal of gene therapy is to insert a normal gene into an organism having a defective gene, in an effort to correct the genetic defect. The polynucleotides disclosed in the present invention offer a means of targeting such genetic defects in a highly accurate manner. Another goal is to insert a new gene that was not present in the host genome, thereby producing a new trait in the host cell. Additional non-limiting examples of gene therapy methods encompassed by the present invention are more thoroughly described elsewhere herein (see, e.g., the sections labeled “Gene Therapy Methods”, and Examples 16, 17 and 18).

The polynucleotides are also useful for identifying individuals from minute biological samples. The United States military, for example, is considering the use of restriction fragment length polymorphism (RFLP) for identification of its personnel. In this technique, an individual's genomic DNA is digested with one or more restriction enzymes, and probed on a Southern blot to yield unique bands for identifying personnel. This method does not suffer from the current limitations of “Dog Tags” which can be lost, switched, or stolen, making positive identification difficult. The polynucleotides of the present invention can be used as additional DNA markers for RFLP.

The polynucleotides of the present invention can also be used as an alternative to RFLP, by determining the actual base-by-base DNA sequence of selected portions of an individual's genome. These sequences can be used to prepare PCR primers for amplifying and isolating such selected DNA, which can then be sequenced. Using this technique, individuals can be identified because each individual will have a unique set of DNA sequences. Once an unique ID database is established for an individual, positive identification of that individual, living or dead, can be made from extremely small tissue samples.

Forensic biology also benefits from using DNA-based identification techniques as disclosed herein. DNA sequences taken from very small biological samples such as tissues, e.g., hair or skin, or body fluids, e.g., blood, saliva, semen, synovial fluid, amniotic fluid, breast milk, lymph, pulmonary sputum or surfactant, urine, fecal matter, etc., can be amplified using PCR. In one prior art technique, gene sequences amplified from polymorphic loci, such as DQa class II HLA gene, are used in forensic biology to identify individuals. (Erlich, H., PCR Technology, Freeman and Co. (1992)). Once these specific polymorphic loci are amplified, they are digested with one or more restriction enzymes, yielding an identifying set of bands on a Southern blot probed with DNA corresponding to the DQa class II HLA gene. Similarly, polynucleotides of the present invention can be used as polymorphic markers for forensic purposes.

There is also a need for reagents capable of identifying the source of a particular tissue. Such need arises, for example, in forensics when presented with tissue of unknown origin. Appropriate reagents can comprise, for example, DNA probes or primers prepared from the sequences of the present invention, specific to tissues, including but not limited to those shown in Table 1B. Panels of such reagents can identify tissue by species and/or by organ type. In a similar fashion, these reagents can be used to screen tissue cultures for contamination. Additional non-limiting examples of such uses are further described herein.

The polynucleotides of the present invention are also useful as hybridization probes for differential identification of the tissue(s) or cell type(s) present in a biological sample. Similarly, polypeptides and antibodies directed to polypeptides of the present invention are useful to provide immunological probes for differential identification of the tissue(s) (e.g., immunohistochemistry assays) or cell type(s) (e.g., immunocytochemistry assays). In addition, for a number of disorders of the above tissues or cells, significantly higher or lower levels of gene expression of the polynucleotides/polypeptides of the present invention may be detected in certain tissues (e.g., tissues expressing polypeptides and/or polynucleotides of the present invention, for example, those disclosed in Table 1B, and/or cancerous and/or wounded tissues) or bodily fluids (e.g., semen, lymph, vaginal pool, serum, plasma, urine, synovial fluid or spinal fluid) taken from an individual having such a disorder, relative to a “standard” gene expression level, i.e., the expression level in healthy tissue from an individual not having the disorder.

Thus, the invention provides a diagnostic method of a disorder, which involves: (a) assaying gene expression level in cells or body fluid of an individual; (b) comparing the gene expression level with a standard gene expression level, whereby an increase or decrease in the assayed gene expression level compared to the standard expression level is indicative of a disorder.

In the very least, the polynucleotides of the present invention can be used as molecular weight markers on Southern gels, as diagnostic probes for the presence of a specific mRNA in a particular cell type, as a probe to “subtract-out” known sequences in the process of discovering novel polynucleotides, for selecting and making oligomers for attachment to a “gene chip” or other support, to raise anti-DNA antibodies using DNA immunization techniques, and as an antigen to elicit an immune response.

Uses of the Polypeptides

Each of the polypeptides identified herein can be used in numerous ways. The following description should be considered exemplary and utilizes known techniques.

Polypeptides and antibodies directed to polypeptides of the present invention are useful to provide immunological probes for differential identification of the tissue(s) (e.g., immunohistochemistry assays such as, for example, ABC immunoperoxidase (Hsu et al., J. Histochem. Cytochem. 29:577-580 (1981)) or cell type(s) (e.g., immunocytochemistry assays).

Antibodies can be used to assay levels of polypeptides encoded by polynucleotides of the invention in a biological sample using classical immunohistological methods known to those of skill in the art (e.g., see Jalkanen, et al., J. Cell. Biol. 101:976-985 (1985); Jalkanen, et al., J. Cell. Biol. 105:3087-3096 (1987)). Other antibody-based methods useful for detecting protein gene expression include immunoassays, such as the enzyme linked immunosorbent assay (ELISA) and the radioimmunoassay (RIA). Suitable antibody assay labels are known in the art and include enzyme labels, such as, glucose oxidase; radioisotopes, such as iodine (¹³¹I, ¹²⁵I, ¹²³I, ¹²¹I), carbon (¹⁴C), sulfur (³⁵S), tritium (³H), indium (^(115m)In, ¹¹³In, ¹¹²In, ¹¹¹In), and technetium (⁹⁹Tc, ^(99m)Tc), thallium, (²⁰¹Ti), gallium (⁶⁸Ga, ⁶⁷Ga), palladium (¹⁰³Pd), molybdenum (⁹⁹Mo), xenon (¹³³Xe), fluorine (¹⁸F) ¹⁵³Sm, ¹⁷⁷Lu, ¹⁵⁹Gd, ¹⁴⁹Pm, ¹⁴⁰La, ¹⁷⁵Yb, ¹⁶⁶Ho, ⁹⁰Y, ⁴⁷Sc, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁴²Pr, ¹⁰⁵Rh, ⁹⁷Ru; luminescent labels, such as luminol; and fluorescent labels, such as fluorescein and rhodamine, and biotin.

In addition to assaying levels of polypeptide of the present invention in a biological sample, proteins can also be detected in vivo by imaging. Antibody labels or markers for in vivo imaging of protein include those detectable by X-radiography, NMR or ESR. For X-radiography, suitable labels include radioisotopes such as barium or cesium, which emit detectable radiation but are not overtly harmful to the subject. Suitable markers for NMR and ESR include those with a detectable characteristic spin, such as deuterium, which may be incorporated into the antibody by labeling of nutrients for the relevant hybridoma.

A protein-specific antibody or antibody fragment which has been labeled with an appropriate detectable imaging moiety, such as a radioisotope (for example, ¹³¹I, ¹¹²In, ^(99m)Tc, (¹³¹I, ¹²⁵I, ¹²³I, ¹²¹I), carbon (¹⁴C), sulfur (³⁵S), tritium (³H), indium (^(115m)In, ^(113m)In, ¹¹²In, ¹¹¹In), and technetium (⁹⁹Tc, ^(99m)Tc), thallium (²⁰¹Ti), gallium (⁶⁸Ga, ⁶⁷Ga), palladium (¹⁰³Pd), molybdenum (⁹⁹Mo), xenon (¹³³Xe), fluorine (¹⁸F, ¹⁵³Sm, ¹⁷⁷Lu, ¹⁵⁹Gd, ¹⁴⁹Pm, ¹⁴⁰La, ¹⁷⁵Yb, ¹⁶⁶Ho, ⁹⁰Y, ⁴⁷Sc, ¹⁸⁶Re, ¹⁸⁸Re, ¹⁴²Pr, ¹⁰⁵Rh, ⁹⁷Ru), a radio-opaque susbstance, or a material detectable by nuclear magnetic resonance, is introduced (for example, parenterally, subcutaneously or intraperitoneally) into the mammal to be examined for immune system disorder. It will be understood in the art that the size of the subject and the imaging system used will determine the quantity of imaging moiety needed to produce diagnostic images. In the case of a radioisotope moiety, for a human subject, the quantity of radioactivity injected will normally range from about 5 to 20 millicuries of ^(99m)Tc. The labeled antibody or antibody fragment will then preferentially accumulate at the location of cells which express the polypeptide encoded by a polynucleotide of the invention. In vivo tumor imaging is described in S. W. Burchiel et al, “Immunopharmacokinetics of Radiolabeled Antibodies and Their Fragments” (Chapter 13 in Tumor Imaging: The Radiochemical Detection of Cancer, S. W. Burchiel and B. A. Rhodes, eds., Masson Publishing Inc. (1982)).

In one embodiment, the invention provides a method for the specific delivery of compositions of the invention to cells by administering polypeptides of the invention (e.g., polypeptides encoded by polynucleotides of the invention and/or antibodies) that are associated with heterologous polypeptides or nucleic acids. In one example, the invention provides a method for delivering a therapeutic protein into the targeted cell. In another example, the invention provides a method for delivering a single stranded nucleic acid (e.g., antisense or ribozymes) or double stranded nucleic acid (e.g., DNA that can integrate into the cell's genome or replicate episomally and that can be transcribed) into the targeted cell.

In another embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention in association with toxins or cytotoxic prodrugs.

By “toxin” is meant one or more compounds that bind and activate endogenous cytotoxic effector systems, radioisotopes, holotoxins, modified toxins, catalytic subunits of toxins, or any molecules or enzymes not normally present in or on the surface of a cell that under defined conditions cause the cell's death. Toxins that may be used according to the methods of the invention include, but are not limited to, radioisotopes known in the art, compounds such as, for example, antibodies (or complement fixing containing portions thereof) that bind an inherent or induced endogenous cytotoxic effector system, thymidine kinase, endonuclease, RNAse, alpha toxin, ricin, abrin, Pseudomonas exotoxin A, diphtheria toxin, saporin, momordin, gelonin, pokeweed antiviral protein, alpha-sarcin and cholera toxin. “Toxin” also includes a cytostatic or cytocidal agent, a therapeutic agent or a radioactive metal ion, e.g., alpha-emitters such as, for example, ²¹³Bi, or other radioisotopes such as, for example, ¹⁰³Pd, ¹³³Xe, ¹³¹I, ⁶⁸Ge, ⁵⁷Co, ⁶⁵Zn, ⁸⁵Sr, ³²P, ³⁵S, ⁹⁰Y, ¹⁵³Gd, ¹⁶⁹Yb, ⁵¹Cr, ⁵⁴Mn, ⁷⁵Se, ¹¹³Sn, ⁹⁰Yttrium, ¹¹⁷Tin, ¹⁸⁶Rhenium, ¹⁶⁶Holmium, and ¹⁸⁸Rhenium; luminescent labels, such as luminol; and fluorescent labels, such as fluorescein and rhodamine, and biotin. In a specific embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention or antibodies of the invention in association with the radioisotope ⁹⁰Y. In another specific embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention or antibodies of the invention in association with the radioisotope ¹¹¹In. In a further specific embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention or antibodies of the invention in association with the radioisotope ¹³¹I.

Techniques known in the art may be applied to label polypeptides of the invention (including antibodies). Such techniques include, but are not limited to, the use of bifunctional conjugating agents (see e.g., U.S. Pat. Nos. 5,756,065; 5,714,631; 5,696,239; 5,652,361; 5,505,931; 5,489,425; 5,435,990; 5,428,139; 5,342,604; 5,274,119; 4,994,560; and 5,808,003; the contents of each of which are hereby incorporated by reference in its entirety).

Thus, the invention provides a diagnostic method of a disorder, which involves (a) assaying the expression level of a polypeptide of the present invention in cells or body fluid of an individual; and (b) comparing the assayed polypeptide expression level with a standard polypeptide expression level, whereby an increase or decrease in the assayed polypeptide expression level compared to the standard expression level is indicative of a disorder. With respect to cancer, the presence of a relatively high amount of transcript in biopsied tissue from an individual may indicate a predisposition for the development of the disease, or may provide a means for detecting the disease prior to the appearance of actual clinical symptoms. A more definitive diagnosis of this type may allow health professionals to employ preventative measures or aggressive treatment earlier thereby preventing the development or further progression of the cancer.

Moreover, polypeptides of the present invention can be used to treat or prevent diseases or conditions such as, for example, neural disorders, immune system disorders, muscular disorders, reproductive disorders, gastrointestinal disorders, pulmonary disorders, cardiovascular disorders, renal disorders, proliferative disorders, and/or cancerous diseases and conditions. For example, patients can be administered a polypeptide of the present invention in an effort to replace absent or decreased levels of the polypeptide (e.g., insulin), to supplement absent or decreased levels of a different polypeptide (e.g., hemoglobin S for hemoglobin B, SOD, catalase, DNA repair proteins), to inhibit the activity of a polypeptide (e.g., an oncogene or tumor supressor), to activate the activity of a polypeptide (e.g., by binding to a receptor), to reduce the activity of a membrane bound receptor by competing with it for free ligand (e.g., soluble TNF receptors used in reducing inflammation), or to bring about a desired response (e.g., blood vessel growth inhibition, enhancement of the immune response to proliferative cells or tissues).

Similarly, antibodies directed to a polypeptide of the present invention can also be used to treat disease (as described supra, and elsewhere herein). For example, administration of an antibody directed to a polypeptide of the present invention can bind, and/or neutralize the polypeptide, and/or reduce overproduction of the polypeptide. Similarly, administration of an antibody can activate the polypeptide, such as by binding to a polypeptide bound to a membrane (receptor).

At the very least, the polypeptides of the present invention can be used as molecular weight markers on SDS-PAGE gels or on molecular sieve gel filtration columns using methods well known to those of skill in the art. Polypeptides can also be used to raise antibodies, which in turn are used to measure protein expression from a recombinant cell, as a way of assessing transformation of the host cell. Moreover, the polypeptides of the present invention can be used to test the biological activities described herein.

Diagnostic Assays

The compounds of the present invention are useful for diagnosis, treatment, prevention and/or prognosis of various disorders in mammals, preferably humans. Such disorders include, but are not limited to, those related to biological activities described in Tables 1D.1 and 1D.2 and, also as described herein under the section heading “Biological Activities”.

For a number of disorders, substantially altered (increased or decreased) levels of gene expression can be detected in tissues, cells or bodily fluids (e.g., sera, plasma, urine, semen, synovial fluid or spinal fluid) taken from an individual having such a disorder, relative to a “standard” gene expression level, that is, the expression level in tissues or bodily fluids from an individual not having the disorder. Thus, the invention provides a diagnostic method useful during diagnosis of a disorder, which involves measuring the expression level of the gene encoding the polypeptide in tissues, cells or body fluid from an individual and comparing the measured gene expression level with a standard gene expression level, whereby an increase or decrease in the gene expression level(s) compared to the standard is indicative of a disorder. These diagnostic assays may be performed in vivo or in vitro, such as, for example, on blood samples, biopsy tissue or autopsy tissue.

The present invention is also useful as a prognostic indicator, whereby patients exhibiting enhanced or depressed gene expression will experience a worse clinical outcome relative to patients expressing the gene at a level nearer the standard level.

In certain embodiments, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to diagnose and/or prognosticate diseases and/or disorders associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1B.2, column 5 (Tissue Distribution Library Code).

By “assaying the expression level of the gene encoding the polypeptide” is intended qualitatively or quantitatively measuring or estimating the level of the polypeptide of the invention or the level of the mRNA encoding the polypeptide of the invention in a first biological sample either directly (e.g., by determining or estimating absolute protein level or mRNA level) or relatively (e.g., by comparing to the polypeptide level or mRNA level in a second biological sample). Preferably, the polypeptide expression level or mRNA level in the first biological sample is measured or estimated and compared to a standard polypeptide level or mRNA level, the standard being taken from a second biological sample obtained from an individual not having the disorder or being determined by averaging levels from a population of individuals not having the disorder. As will be appreciated in the art, once a standard polypeptide level or mRNA level is known, it can be used repeatedly as a standard for comparison.

By “biological sample” is intended any biological sample obtained from an individual, cell line, tissue culture, or other source containing polypeptides of the invention (including portions thereof) or mRNA. As indicated, biological samples include body fluids (such as sera, plasma, urine, synovial fluid and spinal fluid) and tissue sources found to express the full length or fragments thereof of a polypeptide or mRNA. Methods for obtaining tissue biopsies and body fluids from mammals are well known in the art. Where the biological sample is to include mRNA, a tissue biopsy is the preferred source.

Total cellular RNA can be isolated from a biological sample using any suitable technique such as the single-step guanidinium-thiocyanate-phenol-chloroform method described in Chomczynski and Sacchi, Anal. Biochem. 162:156-159 (1987). Levels of mRNA encoding the polypeptides of the invention are then assayed using any appropriate method. These include Northern blot analysis, S1 nuclease mapping, the polymerase chain reaction (PCR), reverse transcription in combination with the polymerase chain reaction (RT-PCR), and reverse transcription in combination with the ligase chain reaction (RT-LCR).

The present invention also relates to diagnostic assays such as quantitative and diagnostic assays for detecting levels of polypeptides of the invention, in a biological sample (e.g., cells and tissues), including determination of normal and abnormal levels of polypeptides. Thus, for instance, a diagnostic assay in accordance with the invention for detecting over-expression of polypeptides of the invention compared to normal control tissue samples may be used to detect the presence of tumors. Assay techniques that can be used to determine levels of a polypeptide, such as a polypeptide of the present invention in a sample derived from a host are well-known to those of skill in the art. Such assay methods include radioimmunoassays, competitive-binding assays, Western Blot analysis and ELISA assays. Assaying polypeptide levels in a biological sample can occur using any art-known method.

Assaying polypeptide levels in a biological sample can occur using antibody-based techniques. For example, polypeptide expression in tissues can be studied with classical immunohistological methods (Jalkanen et al., J. Cell. Biol. 101:976-985 (1985); Jalkanen, M., et al., J. Cell. Biol. 105:3087-3096 (1987)). Other antibody-based methods useful for detecting polypeptide gene expression include immunoassays, such as the enzyme linked immunosorbent assay (ELISA) and the radioimmunoassay (RIA). Suitable antibody assay labels are known in the art and include enzyme labels, such as, glucose oxidase, and radioisotopes, such as iodine (¹²⁵I, ¹²¹I), carbon (¹⁴C), sulfur (³⁵S), tritium (³H), indium (¹¹²In), and technetium (^(99m)Tc), and fluorescent labels, such as fluorescein and rhodamine, and biotin.

The tissue or cell type to be analyzed will generally include those which are known, or suspected, to express the gene of inteest (such as, for example, cancer). The protein isolation methods employed herein may, for example, be such as those described in Harlow and Lane (Harlow, E. and Lane, D., 1988, “Antibodies: A Laboratory Manual”, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.), which is incorporated herein by reference in its entirety. The isolated cells can be derived from cell culture or from a patient. The analysis of cells taken from culture may be a necessary step in the assessment of cells that could be used as part of a cell-based gene therapy technique or, alternatively, to test the effect of compounds on the expression of the gene.

For example, antibodies, or fragments of antibodies, such as those described herein, may be used to quantitatively or qualitatively detect the presence of gene products or conserved variants or peptide fragments thereof. This can be accomplished, for example, by immunofluorescence techniques employing a fluorescently labeled antibody coupled with light microscopic, flow cytometric, or fluorimetric detection.

In a preferred embodiment, antibodies, or fragments of antibodies directed to any one or all of the predicted epitope domains of the polypeptides of the invention (shown in Table 1B) may be used to quantitatively or qualitatively detect the presence of gene products or conserved variants or peptide fragments thereof. This can be accomplished, for example, by immunofluorescence techniques employing a fluorescently labeled antibody coupled with light microscopic, flow cytometric, or fluorimetric detection.

In an additional preferred embodiment, antibodies, or fragments of antibodies directed to a conformational epitope of a polypeptide of the invention may be used to quantitatively or qualitatively detect the presence of gene products or conserved variants or peptide fragments thereof. This can be accomplished, for example, by immunofluorescence techniques employing a fluorescently labeled antibody coupled with light microscopic, flow cytometric, or fluorimetric detection.

The antibodies (or fragments thereof), and/or polypeptides of the present invention may, additionally, be employed histologically, as in immunofluorescence, immunoelectron microscopy or non-immunological assays, for in situ detection of gene products or conserved variants or peptide fragments thereof. In situ detection may be accomplished by removing a histological specimen from a patient, and applying thereto a labeled antibody or polypeptide of the present invention. The antibody (or fragment thereof) or polypeptide is preferably applied by overlaying the labeled antibody (or fragment) onto a biological sample. Through the use of such a procedure, it is possible to determine not only the presence of the gene product, or conserved variants or peptide fragments, or polypeptide binding, but also its distribution in the examined tissue. Using the present invention, those of ordinary skill will readily perceive that any of a wide variety of histological methods (such as staining procedures) can be modified in order to achieve such in situ detection.

Immunoassays and non-immunoassays for gene products or conserved variants or peptide fragments thereof will typically comprise incubating a sample, such as a biological fluid, a tissue extract, freshly harvested cells, or lysates of cells which have been incubated in cell culture, in the presence of a detectably labeled antibody capable of binding gene products or conserved variants or peptide fragments thereof, and detecting the bound antibody by any of a number of techniques well-known in the art.

The biological sample may be brought in contact with and immobilized onto a solid phase support or carrier such as nitrocellulose, or other solid support that is capable of immobilizing cells, cell particles or soluble proteins. The support may then be washed with suitable buffers followed by treatment with the detectably labeled antibody or detectable polypeptide of the invention. The solid phase support may then be washed with the buffer a second time to remove unbound antibody or polypeptide. Optionally the antibody is subsequently labeled. The amount of bound label on solid support may then be detected by conventional means.

By “solid phase support or carrier” is intended any support capable of binding an antigen or an antibody. Well-known supports or carriers include glass, polystyrene, polypropylene, polyethylene, dextran, nylon, amylases, natural and modified celluloses, polyacrylamides, gabbros, and magnetite. The nature of the carrier can be either soluble to some extent or insoluble for the purposes of the present invention. The support material may have virtually any possible structural configuration so long as the coupled molecule is capable of binding to an antigen or antibody. Thus, the support configuration may be spherical, as in a bead, or cylindrical, as in the inside surface of a test tube, or the external surface of a rod. Alternatively, the surface may be flat such as a sheet, test strip, etc. Preferred supports include polystyrene beads. Those skilled in the art will know many other suitable carriers for binding antibody or antigen, or will be able to ascertain the same by use of routine experimentation.

The binding activity of a given lot of antibody or antigen polypeptide may be determined according to well known methods. Those skilled in the art will be able to determine operative and optimal assay conditions for each determination by employing routine experimentation.

In addition to assaying polypeptide levels or polynucleotide levels in a biological sample obtained from an individual, polypeptide or polynucleotide can also be detected in vivo by imaging. For example, in one embodiment of the invention, polypeptides and/or antibodies of the invention are used to image diseased cells, such as neoplasms. In another embodiment, polynucleotides of the invention (e.g., polynucleotides complementary to all or a portion of an mRNA) and/or antibodies (e.g., antibodies directed to any one or a combination of the epitopes of a polypeptide of the invention, antibodies directed to a conformational epitope of a polypeptide of the invention, or antibodies directed to the full length polypeptide expressed on the cell surface of a mammalian cell) are used to image diseased or neoplastic cells.

Antibody labels or markers for in vivo imaging of polypeptides of the invention include those detectable by X-radiography, NMR, MRI, CAT-scans or ESR. For X-radiography, suitable labels include radioisotopes such as barium or cesium, which emit detectable radiation but are not overtly harmful to the subject. Suitable markers for NMR and ESR include those with a detectable characteristic spin, such as deuterium, which may be incorporated into the antibody by labeling of nutrients for the relevant hybridoma. Where in vivo imaging is used to detect enhanced levels of polypeptides for diagnosis in humans, it may be preferable to use human antibodies or “humanized” chimeric monoclonal antibodies. Such antibodies can be produced using techniques described herein or otherwise known in the art. For example methods for producing chimeric antibodies are known in the art. See, for review, Morrison, Science 229:1202 (1985); Oi et al., BioTechniques 4:214 (1986); Cabilly et al., U.S. Pat. No. 4,816,567; Taniguchi et al., EP 171496; Morrison et al., EP 173494; Neuberger et al., WO 8601533; Robinson et al., WO 8702671; Boulianne et al., Nature 312:643 (1984); Neuberger et al., Nature 314:268 (1985).

Additionally, any polypeptides of the invention whose presence can be detected, can be administered. For example, polypeptides of the invention labeled with a radio-opaque or other appropriate compound can be administered and visualized in vivo, as discussed, above for labeled antibodies. Further, such polypeptides can be utilized for in vitro diagnostic procedures.

A polypeptide-specific antibody or antibody fragment that has been labeled with an appropriate detectable imaging moiety, such as a radioisotope (for example, ¹³¹I, ¹¹²In, ^(99m)Tc), a radio-opaque substance, or a material detectable by nuclear magnetic resonance, is introduced (for example, parenterally, subcutaneously or intraperitoneally) into the mammal to be examined for a disorder. It will be understood in the art that the size of the subject and the imaging system used will determine the quantity of imaging moiety needed to produce diagnostic images. In the case of a radioisotope moiety, for a human subject, the quantity of radioactivity injected will normally range from about 5 to 20 millicuries of ^(99m)Tc. The labeled antibody or antibody fragment will then preferentially accumulate at the location of cells which contain the antigenic protein. In vivo tumor imaging is described in S. W. Burchiel et al., “Immunopharmacokinetics of Radiolabeled Antibodies and Their Fragments” (Chapter 13 in Tumor Imaging: The Radiochemical Detection of Cancer, S. W. Burchiel and B. A. Rhodes, eds., Masson Publishing Inc. (1982)).

With respect to antibodies, one of the ways in which an antibody of the present invention can be detectably labeled is by linking the same to a reporter enzyme and using the linked product in an enzyme immunoassay (EIA) (Voller, A., “The Enzyme Linked Immunosorbent Assay (ELISA)”, 1978, Diagnostic Horizons 2:1-7, Microbiological Associates Quarterly Publication, Walkersville, Md.); Voller et al., J. Clin. Pathol. 31:507-520 (1978); Butler, J. E., Meth. Enzymol. 73:482-523 (1981); Maggio, E. (ed.), 1980, Enzyme Immunoassay, CRC Press, Boca Raton, Fla.; Ishikawa, E. et al., (eds.), 1981, Enzyme Immunoassay, Kgaku Shoin, Tokyo). The reporter enzyme which is bound to the antibody will react with an appropriate substrate, preferably a chromogenic substrate, in such a manner as to produce a chemical moiety which can be detected, for example, by spectrophotometric, fluorimetric or by visual means. Reporter enzymes which can be used to detectably label the antibody include, but are not limited to, malate dehydrogenase, staphylococcal nuclease, delta-5-steroid isomerase, yeast alcohol dehydrogenase, alpha-glycerophosphate, dehydrogenase, triose phosphate isomerase, horseradish peroxidase, alkaline phosphatase, asparaginase, glucose oxidase, beta-galactosidase, ribonuclease, urease, catalase, glucose-6-phosphate dehydrogenase, glucoamylase and acetylcholinesterase. Additionally, the detection can be accomplished by colorimetric methods which employ a chromogenic substrate for the reporter enzyme. Detection may also be accomplished by visual comparison of the extent of enzymatic reaction of a substrate in comparison with similarly prepared standards.

Detection may also be accomplished using any of a variety of other immunoassays. For example, by radioactively labeling the antibodies or antibody fragments, it is possible to detect polypeptides through the use of a radioimmunoassay (RIA) (see, for example, Weintraub, B., Principles of Radioimmunoassays, Seventh Training Course on Radioligand Assay Techniques, The Endocrine Society, March, 1986, which is incorporated by reference herein). The radioactive isotope can be detected by means including, but not limited to, a gamma counter, a scintillation counter, or autoradiography.

It is also possible to label the antibody with a fluorescent compound. When the fluorescently labeled antibody is exposed to light of the proper wave length, its presence can then be detected due to fluorescence. Among the most commonly used fluorescent labeling compounds are fluorescein isothiocyanate, rhodamine, phycoerythrin, phycocyanin, allophycocyanin, ophthaldehyde and fluorescamine.

The antibody can also be detectably labeled using fluorescence emitting metals such as ¹⁵²Eu, or others of the lanthanide series. These metals can be attached to the antibody using such metal chelating groups as diethylenetriaminepentacetic acid (DTPA) or ethylenediaminetetraacetic acid (EDTA).

The antibody also can be detectably labeled by coupling it to a chemiluminescent compound. The presence of the chemiluminescent-tagged antibody is then determined by detecting the presence of luminescence that arises during the course of a chemical reaction. Examples of particularly useful chemiluminescent labeling compounds are luminol, isoluminol, theromatic acridinium ester, imidazole, acridinium salt and oxalate ester.

Likewise, a bioluminescent compound may be used to label the antibody of the present invention. Bioluminescence is a type of chemiluminescence found in biological systems in, which a catalytic protein increases the efficiency of the chemiluminescent reaction. The presence of a bioluminescent protein is determined by detecting the presence of luminescence. Important bioluminescent compounds for purposes of labeling are luciferin, luciferase and aequorin.

Methods for Detecting Diseases

In general, a disease may be detected in a patient based on the presence of one or more proteins of the invention and/or polynucleotides encoding such proteins in a biological sample (for example, blood, sera, urine, and/or tumor biopsies) obtained from the patient. In other words, such proteins may be used as markers to indicate the presence or absence of a disease or disorder, including cancer and/or as described elsewhere herein. In addition, such proteins may be useful for the detection of other diseases and cancers. The binding agents provided herein generally permit detection of the level of antigen that binds to the agent in the biological sample. Polynucleotide primers and probes may be used to detect the level of mRNA encoding polypeptides of the invention, which is also indicative of the presence or absence of a disease or disorder, including cancer. In general, polypeptides of the invention should be present at a level that is at least three fold higher in diseased tissue than in normal tissue.

There are a variety of assay formats known to those of ordinary skill in the art for using a binding agent to detect polypeptide markers in a sample. See, e.g., Harlow and Lane, supra. In general, the presence or absence of a disease in a patient may be determined by (a) contacting a biological sample obtained from a patient with a binding agent; (b) detecting in the sample a level of polypeptide that binds to the binding agent; and (c) comparing the level of polypeptide with a predetermined cut-off value.

In a preferred embodiment, the assay involves the use of a binding agent(s) immobilized on a solid support to bind to and remove the polypeptide of the invention from the remainder of the sample. The bound polypeptide may then be detected using a detection reagent that contains a reporter group and specifically binds to the binding agent/polypeptide complex. Such detection reagents may comprise, for example, a binding agent that specifically binds to the polypeptide or an antibody or other agent that specifically binds to the binding agent, such as an anti-immunoglobulin, protein G, protein A or a lectin. Alternatively, a competitive assay may be utilized, in which a polypeptide is labeled with a reporter group and allowed to bind to the immobilized binding agent after incubation of the binding agent with the sample. The extent to which components of the sample inhibit the binding of the labeled polypeptide to the binding agent is indicative of the reactivity of the sample with the immobilized binding agent. Suitable polypeptides for use within such assays include polypeptides of the invention and portions thereof, or antibodies, to which the binding agent binds, as described above.

The solid support may be any material known to those of skill in the art to which polypeptides of the invention may be attached. For example, the solid support may be a test well in a microtiter plate or a nitrocellulose or other suitable membrane. Alternatively, the support may be a bead or disc, such as glass fiberglass, latex or a plastic material such as polystyrene or polyvinylchloride. The support may also be a magnetic particle or a fiber optic sensor, such as those disclosed, for example, in U.S. Pat. No. 5,359,681. The binding agent may be immobilized on the solid support using a variety of techniques known to those of skill in the art, which are amply described in the patent and scientific literature. In the context of the present invention, the term “immobilization” refers to both noncovalent association, such as adsorption, and covalent attachment (which may be a direct linkage between the agent and functional groups on the support or may be a linkage by way of a cross-linking agent). Immobilization by adsorption to a well in a microtiter plate or to a membrane is preferred. In such cases, adsorption may be achieved by contacting the binding agent, in a suitable buffer, with the solid support for the suitable amount of time. The contact time varies with temperature, but is typically between about 1 hour and about 1 day. In general, contacting a well of plastic microtiter plate (such as polystyrene or polyvinylchloride) with an amount of binding agent ranging from about 10 ng to about 10 μg, and preferably about 100 ng to about 1 μg, is sufficient to immobilize an adequate amount of binding agent.

Covalent attachment of binding agent to a solid support may generally be achieved by first reacting the support with a bifunctional reagent that will react with both the support and a functional group, such as a hydroxyl or amino group, on the binding agent. For example, the binding agent may be covalently attached to supports having an appropriate polymer coating using benzoquinone or by condensation of an aldehyde group on the support with an amine and an active hydrogen on the binding partner (see, e.g., Pierce Immunotechnology Catalog and Handbook, 1991, at A12-A13).

Gene Therapy Methods

Also encompassed by the invention are gene therapy methods for treating or preventing disorders, diseases and conditions. The gene therapy methods relate to the introduction of nucleic acid (DNA, RNA and antisense DNA or RNA) sequences into an animal to achieve expression of the polypeptide of the present invention. This method requires a polynucleotide which codes for a polypeptide of the present invention operatively linked to a promoter and any other genetic elements necessary for the expression of the polypeptide by the target tissue. Such gene therapy and delivery techniques are known in the art, see, for example, WO90/11092, which is herein incorporated by reference.

Thus, for example, cells from a patient may be engineered with a polynucleotide (DNA or RNA) comprising a promoter operably linked to a polynucleotide of the present invention ex vivo, with the engineered cells then being provided to a patient to be treated with the polypeptide of the present invention. Such methods are well-known in the art. For example, see Belldegrun, A., et al., J. Natl. Cancer Inst. 85: 207-216 (1993); Ferrantini, M. et al., Cancer Research 53: 1107-1112 (1993); Ferrantini, M. et al., J. Immunology 153: 4604-4615 (1994); Kaido, T., et al., Int. J. Cancer 60: 221-229 (1995); Ogura, H., et al., Cancer Research 50: 5102-5106 (1990); Santodonato, L., et al., Human Gene Therapy 7:1-10 (1996); Santodonato, L., et al., Gene Therapy 4:1246-1255 (1997); and Zhang, J.-F. et al., Cancer Gene Therapy 3: 31-38 (1996)), which are herein incorporated by reference. In one embodiment, the cells which are engineered are arterial cells. The arterial cells may be reintroduced into the patient through direct injection to the artery, the tissues surrounding the artery, or through catheter injection.

As discussed in more detail below, the polynucleotide constructs can be delivered by any method that delivers injectable materials to the cells of an animal, such as, injection into the interstitial space of tissues (heart, muscle, skin, lung, liver, and the like). The polynucleotide constructs may be delivered in a pharmaceutically acceptable liquid or aqueous carrier.

In one embodiment, the polynucleotide of the present invention is delivered as a naked polynucleotide. The term “naked” polynucleotide, DNA or RNA refers to sequences that are free from any delivery vehicle that acts to assist, promote or facilitate entry into the cell, including viral sequences, viral particles, liposome formulations, LIPOFECTIN™ or precipitating agents and the like. However, the polynucleotide of the present invention can also be delivered in liposome formulations and LIPOFECTIN™ formulations and the like can be prepared by methods well known to those skilled in the art. Such methods are described, for example, in U.S. Pat. Nos. 5,593,972, 5,589,466, and 5,580,859, which are herein incorporated by reference.

The polynucleotide vector constructs used in the gene therapy method are preferably constructs that will not integrate into the host genome nor will they contain sequences that allow for replication. Appropriate vectors include pWLNEO, pSV2CAT, pOG44, pXT1 and pSG available from STRATAGENE™; pSVK3, pBPV, pMSG and pSVL available from PHARMACIA™; and pEF1/V5, pcDNA3.1, and pRc/CMV2 available from Invitrogen. Other suitable vectors will be readily apparent to the skilled artisan.

Any strong promoter known to those skilled in the art can be used for driving the expression of the polynucleotide sequence. Suitable promoters include adenoviral promoters, such as the adenoviral major late promoter; or heterologous promoters, such as the cytomegalovirus (CMV) promoter; the respiratory syncytial virus (RSV) promoter; inducible promoters, such as the MMT promoter, the metallothionein promoter; heat shock promoters; the albumin promoter; the ApoAI promoter; human globin promoters; viral thymidine kinase promoters, such as the Herpes Simplex thymidine kinase promoter; retroviral LTRs; the b-actin promoter; and human growth hormone promoters. The promoter also may be the native promoter for the polynucleotide of the present invention.

Unlike other gene therapy techniques, one major advantage of introducing naked nucleic acid sequences into target cells is the transitory nature of the polynucleotide synthesis in the cells. Studies have shown that non-replicating DNA sequences can be introduced into cells to provide production of the desired polypeptide for periods of up to six months.

The polynucleotide construct can be delivered to the interstitial space of tissues within the an animal, including of muscle, skin, brain, lung, liver, spleen, bone marrow, thymus, heart, lymph, blood, bone, cartilage, pancreas, kidney, gall bladder, stomach, intestine, testis, ovary, uterus, rectum, nervous system, eye, gland, and connective tissue. Interstitial space of the tissues comprises the intercellular, fluid, mucopolysaccharide matrix among the reticular fibers of organ tissues, elastic fibers in the walls of vessels or chambers, collagen fibers of fibrous tissues, or that same matrix within connective tissue ensheathing muscle cells or in the lacunae of bone. It is similarly the space occupied by the plasma of the circulation and the lymph fluid of the lymphatic channels. Delivery to the interstitial space of muscle tissue is preferred for the reasons discussed below. They may be conveniently delivered by injection into the tissues comprising these cells. They are preferably delivered to and expressed in persistent, non-dividing cells which are differentiated, although delivery and expression may be achieved in non-differentiated or less completely differentiated cells, such as, for example, stem cells of blood or skin fibroblasts. In vivo muscle cells are particularly competent in their ability to take up and express polynucleotides.

For the naked nucleic acid sequence injection, an effective dosage amount of DNA or RNA will be in the range of from about 0.05 mg/kg body weight to about 50 mg/kg body weight. Preferably the dosage will be from about 0.005 mg/kg to about 20 mg/kg and more preferably from about 0.05 mg/kg to about 5 mg/kg. Of course, as the artisan of ordinary skill will appreciate, this dosage will vary according to the tissue site of injection. The appropriate and effective dosage of nucleic acid sequence can readily be determined by those of ordinary skill in the art and may depend on the condition being treated and the route of administration.

The preferred route of administration is by the parenteral route of injection into the interstitial space of tissues. However, other parenteral routes may also be used, such as, inhalation of an aerosol formulation particularly for delivery to lungs or bronchial tissues, throat or mucous membranes of the nose. In addition, naked DNA constructs can be delivered to arteries during angioplasty by the catheter used in the procedure.

The naked polynucleotides are delivered by any method known in the art, including, but not limited to, direct needle injection at the delivery site, intravenous injection, topical administration, catheter infusion, and so-called “gene guns”. These delivery methods are known in the art.

The constructs may also be delivered with delivery vehicles such as viral sequences, viral particles, liposome formulations, LIPOFECTIN™, precipitating agents, etc. Such methods of delivery are known in the art.

In certain embodiments, the polynucleotide constructs are complexed in a liposome preparation. Liposomal preparations for use in the instant invention include cationic (positively charged), anionic (negatively charged) and neutral preparations. However, cationic liposomes are particularly preferred because a tight charge complex can be formed between the cationic liposome and the polyanionic nucleic acid. Cationic liposomes have been shown to mediate intracellular delivery of plasmid DNA (Feigner et al., Proc. Natl. Acad. Sci. USA (1987) 84:7413-7416, which is herein incorporated by reference); mRNA (Malone et al., Proc. Natl. Acad. Sci. USA (1989) 86:6077-6081, which is herein incorporated by reference); and purified transcription factors (Debs et al., J. Biol. Chem. (1990) 265:10189-10192, which is herein incorporated by reference), in functional form.

Cationic liposomes are readily available. For example, N[1-2,3-dioleyloxy)propyl]-N,N,N-triethylammonium (DOTMA) liposomes are particularly useful and are available under the trademark LIPOFECTINT™, from GIBCO BRL, Grand Island, N.Y. (See, also, Feigner et al., Proc. Natl. Acad. Sci. USA (1987) 84:7413-7416, which is herein incorporated by reference). Other commercially available liposomes include transfectace (DDAB/DOPE) and DOTAP/DOPE (Boehringer).

Other cationic liposomes can be prepared from readily available materials using techniques well known in the art. See, e.g. PCT Publication No. WO 90/11092 (which is herein incorporated by reference) for a description of the synthesis of DOTAP (1,2-bis(oleoyloxy)-3-(trimethylammonio)propane) liposomes. Preparation of DOTMA liposomes is explained in the literature, see, e.g., P. Felgner et al., Proc. Natl. Acad. Sci. USA 84:7413-7417, which is herein incorporated by reference. Similar methods can be used to prepare liposomes from other cationic lipid materials.

Similarly, anionic and neutral liposomes are readily available, such as from Avanti Polar Lipids (Birmingham, Ala.), or can be easily prepared using readily available materials. Such materials include phosphatidyl, choline, cholesterol, phosphatidyl ethanolamine, dioleoylphosphatidyl choline (DOPC), dioleoylphosphatidyl glycerol (DOPG), dioleoylphoshatidyl ethanolamine (DOPE), among others. These materials can also be mixed with the DOTMA and DOTAP starting materials in appropriate ratios. Methods for making liposomes using these materials are well known in the art.

For example, commercially dioleoylphosphatidyl choline (DOPC), dioleoylphosphatidyl glycerol (DOPG), and dioleoylphosphatidyl ethanolamine (DOPE) can be used in various combinations to make conventional liposomes, with or without the addition of cholesterol. Thus, for example, DOPG/DOPC vesicles can be prepared by drying 50 mg each of DOPG and DOPC under a stream of nitrogen gas into a sonication vial. The sample is placed under a vacuum pump overnight and is hydrated the following day with deionized water. The sample is then sonicated for 2 hours in a capped vial, using a Heat Systems model 350 sonicator equipped with an inverted cup (bath type) probe at the maximum setting while the bath is circulated at 15EC. Alternatively, negatively charged vesicles can be prepared without sonication to produce multilamellar vesicles or by extrusion through nucleopore membranes to produce unilamellar vesicles of discrete size. Other methods are known and available to those of skill in the art.

The liposomes can comprise multilamellar vesicles (MLVs), small unilamellar vesicles (SUVs), or large unilamellar vesicles (LUVs), with SUVs being preferred. The various liposome-nucleic acid complexes are prepared using methods well known in the art. See, e.g., Straubinger et al., Methods of Immunology (1983), 101:512-527, which is herein incorporated by reference. For example, MLVs containing nucleic acid can be prepared by depositing a thin film of phospholipid on the walls of a glass tube and subsequently hydrating with a solution of the material to be encapsulated. SUVs are prepared by extended sonication of MLVs to produce a homogeneous population of unilamellar liposomes. The material to be entrapped is added to a suspension of preformed MLVs and then sonicated. When using liposomes containing cationic lipids, the dried lipid film is resuspended in an appropriate solution such as sterile water or an isotonic buffer solution such as 10 mM Tris/NaCl, sonicated, and then the preformed liposomes are mixed directly with the DNA. The liposome and DNA form a very stable complex due to binding of the positively charged liposomes to the cationic DNA. SUVs find use with small nucleic acid fragments. LUVs are prepared by a number of methods, well known in the art. Commonly used methods include Ca²⁺-EDTA chelation (Papahadjopoulos et al., Biochim. Biophys. Acta (1975) 394:483; Wilson et al., Cell 17:77 (1979)); ether injection (Deamer, D. and Bangham, A., Biochim. Biophys. Acta 443:629 (1976); Ostro et al., Biochem. Biophys. Res. Commun. 76:836 (1977); Fraley et al., Proc. Natl. Acad. Sci. USA 76:3348 (1979)); detergent dialysis (Enoch, H. and Strittmatter, P., Proc. Natl. Acad. Sci. USA 76:145 (1979)); and reverse-phase evaporation (REV) (Fraley et al., J. Biol. Chem. 255:10431 (1980); Szoka, F. and Papahadjopoulos, D., Proc. Natl. Acad. Sci. USA 75:145 (1978); Schaefer-Ridder et al., Science 215:166 (1982)), which are herein incorporated by reference.

Generally, the ratio of DNA to liposomes will be from about 10:1 to about 1:10. Preferably, the ration will be from about 5:1 to about 1:5. More preferably, the ration will be about 3:1 to about 1:3. Still more preferably, the ratio will be about 1:1.

U.S. Pat. No. 5,676,954 (which is herein incorporated by reference) reports on the injection of genetic material, complexed with cationic liposomes carriers, into mice. U.S. Pat. Nos. 4,897,355, 4,946,787, 5,049,386, 5,459,127, 5,589,466, 5,693,622, 5,580,859, 5,703,055, and international publication no. WO 94/9469 (which are herein incorporated by reference) provide cationic lipids for use in transfecting DNA into cells and mammals. U.S. Pat. Nos. 5,589,466, 5,693,622, 5,580,859, 5,703,055, and international publication no. WO 94/9469 provide methods for delivering DNA-cationic lipid complexes to mammals.

In certain embodiments, cells are engineered, ex vivo or in vivo, using a retroviral particle containing RNA which comprises a sequence encoding a polypeptide of the present invention. Retroviruses from which the retroviral plasmid vectors may be derived include, but are not limited to, Moloney Murine Leukemia Virus, spleen necrosis virus, Rous sarcoma Virus, Harvey Sarcoma Virus, avian leukosis virus, gibbon ape leukemia virus, human immunodeficiency virus, Myeloproliferative Sarcoma Virus, and mammary tumor virus.

The retroviral plasmid vector is employed to transduce packaging cell lines to form producer cell lines. Examples of packaging cells which may be transfected include, but are not limited to, the PE501, PA317, R-2, R-AM, PA12, T19-14X, VT-19-17-H2, RCRE, RCRIP, GP+E-86, GP+envAm12, and DAN cell lines as described in Miller, Human Gene Therapy 1:5-14 (1990), which is incorporated herein by reference in its entirety. The vector may transduce the packaging cells through any means known in the art. Such means include, but are not limited to, electroporation, the use of liposomes, and CaPO₄ precipitation. In one alternative, the retroviral plasmid vector may be encapsulated into a liposome, or coupled to a lipid, and then administered to a host.

The producer cell line generates infectious retroviral vector particles which include polynucleotide encoding a polypeptide of the present invention. Such retroviral vector particles then may be employed, to transduce eukaryotic cells, either in vitro or in vivo. The transduced eukaryotic cells will express a polypeptide of the present invention.

In certain other embodiments, cells are engineered, ex vivo or in vivo, with polynucleotide contained in an adenovirus vector. Adenovirus can be manipulated such that it encodes and expresses a polypeptide of the present invention, and at the same time is inactivated in terms of its ability to replicate in a normal lytic viral life cycle. Adenovirus expression is achieved without integration of the viral DNA into the host cell chromosome, thereby alleviating concerns about insertional mutagenesis. Furthermore, adenoviruses have been used as live enteric vaccines for many years with an excellent safety profile (Schwartz et al. Am. Rev. Respir. Dis.109:233-238 (1974)). Finally, adenovirus mediated gene transfer has been demonstrated in a number of instances including transfer of alpha-1-antitrypsin and CFTR to the lungs of cotton rats (Rosenfeld, M. A. et al. (1991) Science 252:431-434; Rosenfeld et al., (1992) Cell 68:143-155). Furthermore, extensive studies to attempt to establish adenovirus as a causative agent in human cancer were uniformly negative (Green, M. et al. (1979) Proc. Natl. Acad. Sci. USA 76:6606).

Suitable adenoviral vectors useful in the present invention are described, for example, in Kozarsky and Wilson, Curr. Opin. Genet. Devel. 3:499-503 (1993); Rosenfeld et al., Cell 68:143-155 (1992); Engelhardt et al., Human Genet. Ther. 4:759-769 (1993); Yang et al., Nature Genet. 7:362-369 (1994); Wilson et al., Nature 365:691-692 (1993); and U.S. Pat. No. 5,652,224, which are herein incorporated by reference. For example, the adenovirus vector Ad2 is useful and can be grown in human 293 cells. These cells contain the E1 region of adenovirus and constitutively express E1a and E1b, which complement the defective adenoviruses by providing the products of the genes deleted from the vector. In addition to Ad2, other varieties of adenovirus (e.g., Ad3, Ad5, and Ad7) are also useful in the present invention.

Preferably, the adenoviruses used in the present invention are replication deficient. Replication deficient adenoviruses require the aid of a helper virus and/or packaging cell line to form infectious particles. The resulting virus is capable of infecting cells and can express a polynucleotide of interest that is operably linked to a promoter, but cannot replicate in most cells. Replication deficient adenoviruses may be deleted in one or more of all or a portion of the following genes: E1a, E1b, E3, E4, E2a, or L1 through L5.

In certain other embodiments, the cells are engineered, ex vivo or in vivo, using an adeno-associated virus (AAV). AAVs are naturally occurring defective viruses that require helper viruses to produce infectious particles (Muzyczka, N., Curr. Topics in Microbiol. Immunol 158:97 (1992)). It is also one of the few viruses that may integrate its DNA into non-dividing cells. Vectors containing as little as 300 base pairs of AAV can be packaged and can integrate, but space for exogenous DNA is limited to about 4.5 kb. Methods for producing and using such AAVs are known in the art. See, for example, U.S. Pat. Nos. 5,139,941, 5,173,414, 5,354,678, 5,436,146, 5,474,935, 5,478,745, and 5,589,377.

For example, an appropriate AAV vector for use in the present invention will include all the sequences necessary for DNA replication, encapsidation, and host-cell integration. The polynucleotide construct is inserted into the AAV vector using standard cloning methods, such as those found in Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press (1989). The recombinant AAV vector is then transfected into packaging cells which are infected with a helper virus, using any standard technique, including lipofection, electroporation, calcium phosphate precipitation, etc. Appropriate helper viruses include adenoviruses, cytomegaloviruses, vaccinia viruses, or herpes viruses. Once the packaging cells are transfected and infected, they will produce infectious AAV viral particles which contain the polynucleotide construct. These viral particles are then used to transduce eukaryotic cells, either ex vivo or in vivo. The transduced cells will contain the polynucleotide construct integrated into its genome, and will express a polypeptide of the invention.

Another method of gene therapy involves operably associating heterologous control regions and endogenous polynucleotide sequences (e.g. encoding a polypeptide of the present invention) via homologous recombination (see, e.g., U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication No. WO 96/29411, published Sep. 26, 1996; International Publication No. WO 94/12650, published Aug. 4, 1994; Koller et al., Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); and Zijlstra et al., Nature 342:435-438 (1989), which are herein encorporated by reference. This method involves the activation of a gene which is present in the target cells, but which is not normally expressed in the cells, or is expressed at a lower level than desired.

Polynucleotide constructs are made, using standard techniques known in the art, which contain the promoter with targeting sequences flanking the promoter. Suitable promoters are described herein. The targeting sequence is sufficiently complementary to an endogenous sequence to permit homologous recombination of the promoter-targeting sequence with the endogenous sequence. The targeting sequence will be sufficiently near the 5′ end of the desired endogenous polynucleotide sequence so the promoter will be operably linked to the endogenous sequence upon homologous recombination.

The promoter and the targeting sequences can be amplified using PCR. Preferably, the amplified promoter contains distinct restriction enzyme sites on the 5′ and 3′ ends. Preferably, the 3′ end of the first targeting sequence contains the same restriction enzyme site as the 5′ end of the amplified promoter and the 5′ end of the second targeting sequence contains the same restriction site as the 3′ end of the amplified promoter. The amplified promoter and targeting sequences are digested and ligated together.

The promoter-targeting sequence construct is delivered to the cells, either as naked polynucleotide, or in conjunction with transfection-facilitating agents, such as liposomes, viral sequences, viral particles, whole viruses, lipofection, precipitating agents, etc., described in more detail above. The P promoter-targeting sequence can be delivered by any method, included direct needle injection, intravenous injection, topical administration, catheter infusion, particle accelerators, etc. The methods are described in more detail below.

The promoter-targeting sequence construct is taken up by cells. Homologous recombination between the construct and the endogenous sequence takes place, such that an endogenous sequence is placed under the control of the promoter. The promoter then drives the expression of the endogenous sequence.

The polynucleotide encoding a polypeptide of the present invention may contain a secretory signal sequence that facilitates secretion of the protein. Typically, the signal sequence is positioned in the coding region of the polynucleotide to be expressed towards or at the 5′ end of the coding region. The signal sequence may be homologous or heterologous to the polynucleotide of interest and may be homologous or heterologous to the cells to be transfected. Additionally, the signal sequence may be chemically synthesized using methods known in the art.

Any mode of administration of any of the above-described polynucleotides constructs can be used so long as the mode results in the expression of one or more molecules in an amount sufficient to provide a therapeutic effect. This includes direct needle injection, systemic injection, catheter infusion, biolistic injectors, particle accelerators (i.e., “gene guns”), gelfoam sponge depots, other commercially available depot materials, osmotic pumps (e.g., Alza minipumps), oral or suppositorial solid (tablet or pill) pharmaceutical, and decanting or topical applications during surgery. For example, direct injection of naked calcium phosphate-precipitated plasmid into rat liver and rat spleen or a protein-coated plasmid into the portal vein has resulted in gene expression of the foreign gene in the rat livers (Kaneda et al., Science 243:375 (1989)).

A preferred method of local administration is by direct injection. Preferably, a recombinant molecule of the present invention complexed with a delivery vehicle is administered by direct injection into or locally within the area of arteries. Administration of a composition locally within the area of arteries refers to injecting the composition centimeters and preferably, millimeters within arteries.

Another method of local administration is to contact a polynucleotide construct of the present invention in or around a surgical wound. For example, a patient can undergo surgery and the polynucleotide construct can be coated on the surface of tissue inside the wound or the construct can be injected into areas of tissue inside the wound.

Therapeutic compositions useful in systemic administration include recombinant molecules of the present invention complexed to a targeted delivery vehicle of the present invention. Suitable delivery vehicles for use with systemic administration comprise liposomes comprising ligands for targeting the vehicle to a particular site. In specific embodiments, suitable delivery vehicles for use with systemic administration comprise liposomes comprising polypeptides of the invention for targeting the vehicle to a particular site.

Preferred methods of systemic administration, include intravenous injection, aerosol, oral and percutaneous (topical) delivery. Intravenous injections can be performed using methods standard in the art. Aerosol delivery can also be performed using methods standard in the art (see, for example, Stribling et al., Proc. Natl. Acad. Sci. USA 189:11277-11281, 1992, which is incorporated herein by reference). Oral delivery can be performed by complexing a polynucleotide construct of the present invention to a carrier capable of withstanding degradation by digestive enzymes in the gut of an animal. Examples of such carriers, include plastic capsules or tablets, such as those known in the art. Topical delivery can be performed by mixing a polynucleotide construct of the present invention with a lipophilic reagent (e.g., DMSO) that is capable of passing into the skin.

Determining an effective amount of substance to be delivered can depend upon a number of factors including, for example, the chemical structure and biological activity of the substance, the age and weight of the animal, the precise condition requiring treatment and its severity, and the route of administration. The frequency of treatments depends upon a number of factors, such as the amount of polynucleotide constructs administered per dose, as well as the health and history of the subject. The precise amount, number of doses, and timing of doses will be determined by the attending physician or veterinarian.

Therapeutic compositions of the present invention can be administered to any animal, preferably to mammals and birds. Preferred mammals include humans, dogs, cats, mice, rats, rabbits sheep, cattle, horses and pigs, with humans being particularly preferred.

Biological Activities

Polynucleotides or polypeptides, or agonists or antagonists of the present invention, can be used in assays to test for one or more biological activities. If these polynucleotides or polypeptides, or agonists or antagonists of the present invention, do exhibit activity in a particular assay, it is likely that these molecules may be involved in the diseases associated with the biological activity. Thus, the polynucleotides and polypeptides, and agonists or antagonists could be used to treat the associated disease.

Members of the secreted family of proteins are believed to be involved in biological activities associated with, for example, cellular signaling. Accordingly, compositions of the invention (including polynucleotides, polypeptides and antibodies of the invention, and fragments and variants thereof) may be used in diagnosis, prognosis, prevention and/or treatment of diseases and/or disorders associated with aberrant activity of secreted polypeptides.

In preferred embodiments, compositions of the invention (including polynucleotides, polypeptides and antibodies of the invention, and fragments and variants thereof) may be used in the diagnosis, prognosis, prevention, treatment, and/or amelioration of diseases and/or disorders relating to the gastrointestinal system (e.g., Crohn's disease, pancreatitis, gallstones, antibiotic-associated colitis, duodenitis, gastrointestinal neoplasms, and as described in the “Gastrointestinal Disorders” section below).

In certain embodiments, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to diagnose and/or prognosticate diseases and/or disorders associated with the tissue(s) in which the polypeptide of the invention is expressed including one, two, three, four, five, or more tissues disclosed in Table 1B.2, column 5 (Tissue Distribution Library Code).

Thus, polynucleotides, translation products and antibodies of the invention are useful in the diagnosis, detection, prevention, prognistication, and/or treatment of diseases and/or disorders associated with activities that include, but are not limited to, prohormone activation, neurotransmitter activity, cellular signaling, cellular proliferation, cellular differentiation, and cell migration.

More generally, polynucleotides, translation products and antibodies corresponding to this gene may be useful for the diagnosis, prognosis, prevention, treatment and/or amelioration of diseases and/or disorders associated with the following system or systems.

Immune Activity

Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in preventing, diagnosing, prognosticating, treating, and/or ameliorating diseases, disorders, and/or conditions of the immune system, by, for example, activating or inhibiting the proliferation, differentiation, or mobilization (chemotaxis) of immune cells Immune cells develop through a process called hematopoiesis, producing myeloid (platelets, red blood cells, neutrophils, and macrophages) and lymphoid (B and T lymphocytes) cells from pluripotent stem cells. The etiology of these immune diseases, disorders, and/or conditions may be genetic, somatic, such as cancer and some autoimmune diseases, acquired (e.g., by chemotherapy or toxins), or infectious. Moreover, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention can be used as a marker or detector of a particular immune system disease or disorder.

In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to treat diseases and disorders of the immune system and/or to inhibit or enhance an immune response generated by cells associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1B.2, column 5 (Tissue Distribution Library Code).

Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in preventing, diagnosing, prognosticating, treating and/or ameliorating immunodeficiencies, including both congenital and acquired immunodeficiencies. Examples of B cell immunodeficiencies in which immunoglobulin levels B cell function and/or B cell numbers are decreased include: X-linked agammaglobulinemia (Bruton's disease), X-linked infantile agammaglobulinemia, X-linked immunodeficiency with hyper IgM, non X-linked immunodeficiency with hyper IgM, X-linked lymphoproliferative syndrome (XLP), agammaglobulinemia including congenital and acquired agammaglobulinemia, adult onset agammaglobulinemia, late-onset agammaglobulinemia, dysgammaglobulinemia, hypogammaglobulinemia, unspecified hypogammaglobulinemia, recessive agammaglobulinemia (Swiss type), Selective IgM deficiency, selective IgA deficiency, selective IgG subclass deficiencies, IgG subclass deficiency (with or without IgA deficiency), Ig deficiency with increased IgM, IgG and IgA deficiency with increased IgM, antibody deficiency with normal or elevated Igs, Ig heavy chain deletions, kappa chain deficiency, B cell lymphoproliferative disorder (BLPD), common variable immunodeficiency (CVID), common variable immunodeficiency (CVI) (acquired), and transient hypogammaglobulinemia of infancy.

In specific embodiments, ataxia-telangiectasia or conditions associated with ataxia-telangiectasia are detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated using the polypeptides or polynucleotides of the invention, and/or agonists or antagonists thereof.

Examples of congenital immunodeficiencies in which T cell and/or B cell function and/or number is decreased include, but are not limited to: DiGeorge anomaly, severe combined immunodeficiencies (SCID) (including, but not limited to, X-linked SCID, autosomal recessive SCID, adenosine deaminase deficiency, purine nucleoside phosphorylase (PNP) deficiency, Class II MHC deficiency (Bare lymphocyte syndrome), Wiskott-Aldrich syndrome, and ataxia telangiectasia), thymic hypoplasia, third and fourth pharyngeal pouch syndrome, 22q11.2 deletion, chronic mucocutaneous candidiasis, natural killer cell deficiency (NK), idiopathic CD4+T-lymphocytopenia, immunodeficiency with predominant T cell defect (unspecified), and unspecified immunodeficiency of cell mediated immunity

In specific embodiments, DiGeorge anomaly or conditions associated with DiGeorge anomaly are prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polypeptides or polynucleotides of the invention, or antagonists or agonists thereof.

Other immunodeficiencies that may be prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polypeptides or polynucleotides of the invention, and/or agonists or antagonists thereof, include, but are not limited to, chronic granulomatous disease, Chediak-Higashi syndrome, myeloperoxidase deficiency, leukocyte glucose-6-phosphate dehydrogenase deficiency, X-linked lymphoproliferative syndrome (XLP), leukocyte adhesion deficiency, complement component deficiencies (including C1, C2, C3, C4, C5, C6, C7, C8 and/or C9 deficiencies), reticular dysgenesis, thymic alymphoplasia-aplasia, immunodeficiency with thymoma, severe congenital leukopenia, dysplasia with immunodeficiency, neonatal neutropenia, short limbed dwarfism, and Nezelof syndrome-combined immunodeficiency with Igs.

In a preferred embodiment, the immunodeficiencies and/or conditions associated with the immunodeficiencies recited above are prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

In a preferred embodiment polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used as an agent to boost immunoresponsiveness among immunodeficient individuals. In specific embodiments, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used as an agent to boost immunoresponsiveness among B cell and/or T cell immunodeficient individuals.

The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in preventing, detecting, diagnosing, prognosticating, treating and/or ameliorating autoimmune disorders. Many autoimmune disorders result from inappropriate recognition of self as foreign material by immune cells. This inappropriate recognition results in an immune response leading to the destruction of the host tissue. Therefore, the administration of polynucleotides and polypeptides of the invention that can inhibit an immune response, particularly the proliferation, differentiation, or chemotaxis of T-cells, may be an effective therapy in preventing autoimmune disorders.

Autoimmune diseases or disorders that may be prevented, detected, diagnosed, prognosticated, treated, and/or ameliorated by polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include, but are not limited to, one or more of the following: systemic lupus erythematosus, rheumatoid arthritis, ankylosing spondylitis, multiple sclerosis, autoimmune thyroiditis, Hashimoto's thyroiditis, autoimmune hemolytic anemia, hemolytic anemia, thrombocytopenia, autoimmune thrombocytopenia purpura, autoimmune neonatal thrombocytopenia, idiopathic thrombocytopenia purpura, purpura (e.g., Henloch-Scoenlein purpura), autoimmunocytopenia, Goodpasture's syndrome, Pemphigus vulgaris, myasthenia gravis, Grave's disease (hyperthyroidism), and insulin-resistant diabetes mellitus.

Additional disorders that are likely to have an autoimmune component that may be prevented, detected, diagnosed, prognosticated, treated and/or ameliorated with the compositions of the invention include, but are not limited to, type II collagen-induced arthritis, antiphospholipid syndrome, dermatitis, allergic encephalomyelitis, myocarditis, relapsing polychondritis, rheumatic heart disease, neuritis, uveitis ophthalmia, polyendocrinopathies, Reiter's Disease, Stiff-Man Syndrome, autoimmune pulmonary inflammation, autism, Guillain-Barre Syndrome, insulin dependent diabetes mellitus, and autoimmune inflammatory eye disorders.

Additional disorders that are likely to have an autoimmune component that may be prevented, detected, diagnosed, prognosticated, treated and/or ameliorated with the compositions of the invention include, but are not limited to, scleroderma with anti-collagen antibodies (often characterized, e.g., by nucleolar and other nuclear antibodies), mixed connective tissue disease (often characterized, e.g., by antibodies to extractable nuclear antigens (e.g., ribonucleoprotein)), polymyositis (often characterized, e.g., by nonhistone ANA), pernicious anemia (often characterized, e.g., by antiparietal cell, microsomes, and intrinsic factor antibodies), idiopathic Addison's disease (often characterized, e.g., by humoral and cell-mediated adrenal cytotoxicity, infertility (often characterized, e.g., by antispermatozoal antibodies), glomerulonephritis (often characterized, e.g., by glomerular basement membrane antibodies or immune complexes), bullous pemphigoid (often characterized, e.g., by IgG and complement in basement membrane), Sjogren's syndrome (often characterized, e.g., by multiple tissue antibodies, and/or a specific nonhistone ANA (SS-B)), diabetes mellitus (often characterized, e.g., by cell-mediated and humoral islet cell antibodies), and adrenergic drug resistance (including adrenergic drug resistance with asthma or cystic fibrosis) (often characterized, e.g., by beta-adrenergic receptor antibodies).

Additional disorders that may have an autoimmune component that may be prevented, detected, diagnosed, prognosticated, treated and/or ameliorated with the compositions of the invention include, but are not limited to, chronic active hepatitis (often characterized, e.g., by smooth muscle antibodies), primary biliary cirrhosis (often characterized, e.g., by mitochondria antibodies), other endocrine gland failure (often characterized, e.g., by specific tissue antibodies in some cases), vitiligo (often characterized, e.g., by melanocyte antibodies), vasculitis (often characterized, e.g., by Ig and complement in vessel walls and/or low serum complement), post-MI (often characterized, e.g., by myocardial antibodies), cardiotomy syndrome (often characterized, e.g., by myocardial antibodies), urticaria (often characterized, e.g., by IgG and IgM antibodies to IgE), atopic dermatitis (often characterized, e.g., by IgG and IgM antibodies to IgE), asthma (often characterized, e.g., by IgG and IgM antibodies to IgE), and many other inflammatory, granulomatous, degenerative, and atrophic disorders.

In a preferred embodiment, the autoimmune diseases and disorders and/or conditions associated with the diseases and disorders recited above are prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using for example, antagonists or agonists, polypeptides or polynucleotides, or antibodies of the present invention. In a specific preferred embodiment, rheumatoid arthritis is prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

In another specific preferred embodiment, systemic lupus erythematosus is prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention. In another specific preferred embodiment, idiopathic thrombocytopenia purpura is prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

In another specific preferred embodiment IgA nephropathy is prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

In a preferred embodiment, the autoimmune diseases and disorders and/or conditions associated with the diseases and disorders recited above are prevented, detected, diagnosed, prognosticated, treated and/or ameliorated using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention

In preferred embodiments, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a immunosuppressive agent(s).

Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating diseases, disorders, and/or conditions of hematopoietic cells. Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used to increase differentiation and proliferation of hematopoietic cells, including the pluripotent stem cells, in an effort to treat or prevent those diseases, disorders, and/or conditions associated with a decrease in certain (or many) types hematopoietic cells, including but not limited to, leukopenia, neutropenia, anemia, and thrombocytopenia. Alternatively, Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used to increase differentiation and proliferation of hematopoietic cells, including the pluripotent stem cells, in an effort to treat or prevent those diseases, disorders, and/or conditions associated with an increase in certain (or many) types of hematopoietic cells, including but not limited to, histiocytosis.

Allergic reactions and conditions, such as asthma (particularly allergic asthma) or other respiratory problems, may also be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated using polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof. Moreover, these molecules can be used to treat, prevent, prognose, and/or diagnose anaphylaxis, hypersensitivity to an antigenic molecule, or blood group incompatibility.

Additionally, polypeptides or polynucleotides of the invention, and/or agonists or antagonists thereof, may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate IgE-mediated allergic reactions. Such allergic reactions include, but are not limited to, asthma, rhinitis, and eczema. In specific embodiments, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to modulate IgE concentrations in vitro or in vivo.

Moreover, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention have uses in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of inflammatory conditions. For example, since polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists of the invention may inhibit the activation, proliferation and/or differentiation of cells involved in an inflammatory response, these molecules can be used to prevent and/or treat chronic and acute inflammatory conditions. Such inflammatory conditions include, but are not limited to, for example, inflammation associated with infection (e.g., septic shock, sepsis, or systemic inflammatory response syndrome), ischemia-reperfusion injury, endotoxin lethality, complement-mediated hyperacute rejection, nephritis, cytokine or chemokine induced lung injury, inflammatory bowel disease, Crohn's disease, over production of cytokines (e.g., TNF or IL-1.), respiratory disorders (e.g., asthma and allergy); gastrointestinal disorders (e.g., inflammatory bowel disease); cancers (e.g., gastric, ovarian, lung, bladder, liver, and breast); CNS disorders (e.g., multiple sclerosis; ischemic brain injury and/or stroke, traumatic brain injury, neurodegenerative disorders (e.g., Parkinson's disease and Alzheimer's disease); AIDS-related dementia; and prion disease); cardiovascular disorders (e.g., atherosclerosis, myocarditis, cardiovascular disease, and cardiopulmonary bypass complications); as well as many additional diseases, conditions, and disorders that are characterized by inflammation (e.g., hepatitis, rheumatoid arthritis, gout, trauma, pancreatitis, sarcoidosis, dermatitis, renal ischemia-reperfusion injury, Grave's disease, systemic lupus erythematosus, diabetes mellitus, and allogenic transplant rejection).

Because inflammation is a fundamental defense mechanism, inflammatory disorders can effect virtually any tissue of the body. Accordingly, polynucleotides, polypeptides, and antibodies of the invention, as well as agonists or antagonists thereof, have uses in the treatment of tissue-specific inflammatory disorders, including, but not limited to, adrenalitis, alveolitis, angiocholecystitis, appendicitis, balanitis, blepharitis, bronchitis, bursitis, carditis, cellulitis, cervicitis, cholecystitis, chorditis, cochlitis, colitis, conjunctivitis, cystitis, dermatitis, diverticulitis, encephalitis, endocarditis, esophagitis, eustachitis, fibrositis, folliculitis, gastritis, gastroenteritis, gingivitis, glossitis, hepatosplenitis, keratitis, labyrinthitis, laryngitis, lymphangitis, mastitis, media otitis, meningitis, metritis, mucitis, myocarditis, myosititis, myringitis, nephritis, neuritis, orchitis, osteochondritis, otitis, pericarditis, peritendonitis, peritonitis, pharyngitis, phlebitis, poliomyelitis, prostatitis, pulpitis, retinitis, rhinitis, salpingitis, scleritis, sclerochoroiditis, scrotitis, sinusitis, spondylitis, steatitis, stomatitis, synovitis, syringitis, tendonitis, tonsillitis, urethritis, and vaginitis.

In specific embodiments, polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, are useful to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate organ transplant rejections and graft-versus-host disease. Organ rejection occurs by host immune cell destruction of the transplanted tissue through an immune response. Similarly, an immune response is also involved in GVHD, but, in this case, the foreign transplanted immune cells destroy the host tissues. Polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, that inhibit an immune response, particularly the activation, proliferation, differentiation, or chemotaxis of T-cells, may be an effective therapy in preventing organ rejection or GVHD. In specific embodiments, polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, that inhibit an immune response, particularly the activation, proliferation, differentiation, or chemotaxis of T-cells, may be an effective therapy in preventing experimental allergic and hyperacute xenograft rejection.

In other embodiments, polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, are useful to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate immune complex diseases, including, but not limited to, serum sickness, post streptococcal glomerulonephritis, polyarteritis nodosa, and immune complex-induced vasculitis.

Polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the invention can be used to treat, detect, and/or prevent infectious agents. For example, by increasing the immune response, particularly increasing the proliferation activation and/or differentiation of B and/or T cells, infectious diseases may be treated, detected, and/or prevented. The immune response may be increased by either enhancing an existing immune response, or by initiating a new immune response. Alternatively, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may also directly inhibit the infectious agent (refer to section of application listing infectious agents, etc), without necessarily eliciting an immune response.

In another embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a vaccine adjuvant that enhances immune responsiveness to an antigen. In a specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance tumor-specific immune responses.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance anti-viral immune responses. Anti-viral immune responses that may be enhanced using the compositions of the invention as an adjuvant, include virus and virus associated diseases or symptoms described herein or otherwise known in the art. In specific embodiments, the compositions of the invention are used as an adjuvant to enhance an immune response to a virus, disease, or symptom selected from the group consisting of: AIDS, meningitis, Dengue, EBV, and hepatitis (e.g., hepatitis B). In another specific embodiment, the compositions of the invention are used as an adjuvant to enhance an immune response to a virus, disease, or symptom selected from the group consisting of: HIV/AIDS, respiratory syncytial virus, Dengue, rotavirus, Japanese B encephalitis, influenza A and B, parainfluenza, measles, cytomegalovirus, rabies, Junin, Chikungunya, Rift Valley Fever, herpes simplex, and yellow fever.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance anti-bacterial or anti-fungal immune responses. Anti-bacterial or anti-fungal immune responses that may be enhanced using the compositions of the invention as an adjuvant, include bacteria or fungus and bacteria or fungus associated diseases or symptoms described herein or otherwise known in the art. In specific embodiments, the compositions of the invention are used as an adjuvant to enhance an immune response to a bacteria or fungus, disease, or symptom selected from the group consisting of: tetanus, Diphtheria, botulism, and meningitis type B.

In another specific embodiment, the compositions of the invention are used as an adjuvant to enhance an immune response to a bacteria or fungus, disease, or symptom selected from the group consisting of: Vibrio cholerae, Mycobacterium leprae, Salmonella typhi, Salmonella paratyphi, Meisseria meningitidis, Streptococcus pneumoniae, Group B streptococcus, Shigella spp., Enterotoxigenic Escherichia coli, Enterohemorrhagic E. coli, and Borrelia burgdorferi.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance anti-parasitic immune responses. Anti-parasitic immune responses that may be enhanced using the compositions of the invention as an adjuvant, include parasite and parasite associated diseases or symptoms described herein or otherwise known in the art. In specific embodiments, the compositions of the invention are used as an adjuvant to enhance an immune response to a parasite. In another specific embodiment, the compositions of the invention are used as an adjuvant to enhance an immune response to Plasmodium (malaria) or Leishmania.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may also be employed to treat infectious diseases including silicosis, sarcoidosis, and idiopathic pulmonary fibrosis; for example, by preventing the recruitment and activation of mononuclear phagocytes.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an antigen for the generation of antibodies to inhibit or enhance immune mediated responses against polypeptides of the invention.

In one embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are administered to an animal (e.g., mouse, rat, rabbit, hamster, guinea pig, pigs, micro-pig, chicken, camel, goat, horse, cow, sheep, dog, cat, non-human primate, and human, most preferably human) to boost the immune system to produce increased quantities of one or more antibodies (e.g., IgG, IgA, IgM, and IgE), to induce higher affinity antibody production and immunoglobulin class switching (e.g., IgG, IgA, IgM, and IgE), and/or to increase an immune response.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a stimulator of B cell responsiveness to pathogens.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an activator of T cells.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent that elevates the immune status of an individual prior to their receipt of immunosuppressive therapies.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to induce higher affinity antibodies.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to increase serum immunoglobulin concentrations.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to accelerate recovery of immunocompromised individuals.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to boost immunoresponsiveness among aged populations and/or neonates.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an immune system enhancer prior to, during, or after bone marrow transplant and/or other transplants (e.g., allogeneic or xenogeneic organ transplantation). With respect to transplantation, compositions of the invention may be administered prior to, concomitant with, and/or after transplantation. In a specific embodiment, compositions of the invention are administered after transplantation, prior to the beginning of recovery of T-cell populations. In another specific embodiment, compositions of the invention are first administered after transplantation after the beginning of recovery of T cell populations, but prior to full recovery of B cell populations.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to boost immunoresponsiveness among individuals having an acquired loss of B cell function. Conditions resulting in an acquired loss of B cell function that may be ameliorated or treated by administering the polypeptides, antibodies, polynucleotides and/or agonists or antagonists thereof, include, but are not limited to, HW Infection, AIDS, bone marrow transplant, and B cell chronic lymphocytic leukemia (CLL).

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to boost immunoresponsiveness among individuals having a temporary immune deficiency. Conditions resulting in a temporary immune deficiency that may be ameliorated or treated by administering the polypeptides, antibodies, polynucleotides and/or agonists or antagonists thereof, include, but are not limited to, recovery from viral infections (e.g., influenza), conditions associated with malnutrition, recovery from infectious mononucleosis, or conditions associated with stress, recovery from measles, recovery from blood transfusion, and recovery from surgery.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a regulator of antigen presentation by monocytes, dendritic cells, and/or B-cells. In one embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention enhance antigen presentation or antagonizes antigen presentation in vitro or in vivo. Moreover, in related embodiments, said enhancement or antagonism of antigen presentation may be useful as an anti-tumor treatment or to modulate the immune system.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to direct an individual's immune system towards development of a humoral response (i.e. TH2) as opposed to a TH1 cellular response.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means to induce tumor proliferation and thus make it more susceptible to anti-neoplastic agents. For example, multiple myeloma is a slowly dividing disease and is thus refractory to virtually all anti-neoplastic regimens. If these cells were forced to proliferate more rapidly their susceptibility profile would likely change.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a stimulator of B cell production in pathologies such as AIDS, chronic lymphocyte disorder and/or Common Variable Immunodificiency.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for generation and/or regeneration of lymphoid tissues following surgery, trauma or genetic defect. In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used in the pretreatment of bone marrow samples prior to transplant.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a gene-based therapy for genetically inherited disorders resulting in immuno-incompetence/immunodeficiency such as observed among SCID patients.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of activating monocytes/macrophages to defend against parasitic diseases that effect monocytes such as Leishmania.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of regulating secreted cytokines that are elicited by polypeptides of the invention.

In another embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used in one or more of the applications described herein, as they may apply to veterinary medicine.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of blocking various aspects of immune responses to foreign agents or self. Examples of diseases or conditions in which blocking of certain aspects of immune responses may be desired include autoimmune disorders such as lupus, and arthritis, as well as immunoresponsiveness to skin allergies, inflammation, bowel disease, injury and diseases/disorders associated with pathogens.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for preventing the B cell proliferation and Ig secretion associated with autoimmune diseases such as idiopathic thrombocytopenic purpura, systemic lupus erythematosus and multiple sclerosis.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a inhibitor of B and/or T cell migration in endothelial cells. This activity disrupts tissue architecture or cognate responses and is useful, for example in disrupting immune responses, and blocking sepsis.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for chronic hypergammaglobulinemia evident in such diseases as monoclonal gammopathy of undetermined significance (MGUS), Waldenstrom's disease, related idiopathic monoclonal gammopathies, and plasmacytomas.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be employed for instance to inhibit polypeptide chemotaxis and activation of macrophages and their precursors, and of neutrophils, basophils, B lymphocytes and some T-cell subsets, e.g., activated and CD8 cytotoxic T cells and natural killer cells, in certain autoimmune and chronic inflammatory and infective diseases. Examples of autoimmune diseases are described herein and include multiple sclerosis, and insulin-dependent diabetes.

The polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may also be employed to treat idiopathic hyper-eosinophilic syndrome by, for example, preventing eosinophil production and migration.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used to enhance or inhibit complement mediated cell lysis.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used to enhance or inhibit antibody dependent cellular cytotoxicity.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may also be employed for treating atherosclerosis, for example, by preventing monocyte infiltration in the artery wall.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be employed to treat adult respiratory distress syndrome (ARDS).

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be useful for stimulating wound and tissue repair, stimulating angiogenesis, and/or stimulating the repair of vascular or lymphatic diseases or disorders. Additionally, agonists and antagonists of the invention may be used to stimulate the regeneration of mucosal surfaces.

In a specific embodiment, polynucleotides or polypeptides, and/or agonists thereof are used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate a disorder characterized by primary or acquired immunodeficiency, deficient serum immunoglobulin production, recurrent infections, and/or immune system dysfunction. Moreover, polynucleotides or polypeptides, and/or agonists thereof may be used to treat or prevent infections of the joints, bones, skin, and/or parotid glands, blood-borne infections (e.g., sepsis, meningitis, septic arthritis, and/or osteomyelitis), autoimmune diseases (e.g., those disclosed herein), inflammatory disorders, and malignancies, and/or any disease or disorder or condition associated with these infections, diseases, disorders and/or malignancies) including, but not limited to, CVID, other primary immune deficiencies, HIV disease, CLL, recurrent bronchitis, sinusitis, otitis media, conjunctivitis, pneumonia, hepatitis, meningitis, herpes zoster (e.g., severe herpes zoster), and/or pneumocystis carnii. Other diseases and disorders that may be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with polynucleotides or polypeptides, and/or agonists of the present invention include, but are not limited to, HW infection, HTLV-BLV infection, lymphopenia, phagocyte bactericidal dysfunction anemia, thrombocytopenia, and hemoglobinuria.

In another embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention are used to treat, and/or diagnose an individual having common variable immunodeficiency disease (“CVID”; also known as “acquired agammaglobulinemia” and “acquired hypogammaglobulinemia”) or a subset of this disease.

In a specific embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate cancers or neoplasms including immune cell or immune tissue-related cancers or neoplasms. Examples of cancers or neoplasms that may be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include, but are not limited to, acute myelogenous leukemia, chronic myelogenous leukemia, Hodgkin's disease, non-Hodgkin's lymphoma, acute lymphocytic anemia (ALL) Chronic lymphocyte leukemia, plasmacytomas, multiple myeloma, Burkitt's lymphoma, EBV-transformed diseases, and/or diseases and disorders described in the section entitled “Hyperproliferative Disorders” elsewhere herein.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for decreasing cellular proliferation of Large B-cell Lymphomas.

In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of decreasing the involvement of B cells and Ig associated with Chronic Myelogenous Leukemia.

In specific embodiments, the compositions of the invention are used as an agent to boost immunoresponsiveness among B cell immunodeficient individuals, such as, for example, an individual who has undergone a partial or complete splenectomy.

Antagonists of the invention include, for example, binding and/or inhibitory antibodies, antisense nucleic acids, ribozymes or soluble forms of the polypeptides of the present invention (e.g., Fc fusion protein; see, e.g., Example 9). Agonists of the invention include, for example, binding or stimulatory antibodies, and soluble forms of the polypeptides (e.g., Fc fusion proteins; see, e.g., Example 9). polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be employed in a composition with a pharmaceutically acceptable carrier, e.g., as described herein.

In another embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are administered to an animal (including, but not limited to, those listed above, and also including transgenic animals) incapable of producing functional endogenous antibody molecules or having an otherwise compromised endogenous immune system, but which is capable of producing human immunoglobulin molecules by means of a reconstituted or partially reconstituted immune system from another animal (see, e.g., published PCT Application Nos. WO98/24893, WO/9634096, WO/9633735, and WO/9110741). Administration of polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention to such animals is useful for the generation of monoclonal antibodies against the polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention.

Blood-Related Disorders

The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to modulate hemostatic (the stopping of bleeding) or thrombolytic (clot dissolving) activity. For example, by increasing hemostatic or thrombolytic activity, polynucleotides or polypeptides, and/or agonists or antagonists of the present invention could be used to treat or prevent blood coagulation diseases, disorders, and/or conditions (e.g., afibrinogenemia, factor deficiencies, hemophilia), blood platelet diseases, disorders, and/or conditions (e.g., thrombocytopenia), or wounds resulting from trauma, surgery, or other causes. Alternatively, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention that can decrease hemostatic or thrombolytic activity could be used to inhibit or dissolve clotting. These molecules could be important in the treatment or prevention of heart attacks (infarction), strokes, or scarring.

In specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate thrombosis, arterial thrombosis, venous thrombosis, thromboembolism, pulmonary embolism, atherosclerosis, myocardial infarction, transient ischemic attack, unstable angina. In specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used for the prevention of occulsion of saphenous grafts, for reducing the risk of periprocedural thrombosis as might accompany angioplasty procedures, for reducing the risk of stroke in patients with atrial fibrillation including nonrheumatic atrial fibrillation, for reducing the risk of embolism associated with mechanical heart valves and or mitral valves disease. Other uses for the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention, include, but are not limited to, the prevention of occlusions in extrcorporeal devices (e.g., intravascular canulas, vascular access shunts in hemodialysis patients, hemodialysis machines, and cardiopulmonary bypass machines).

In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate diseases and disorders of the blood and/or blood forming organs associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1B.2, column 5 (Tissue Distribution Library Code).

The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to modulate hematopoietic activity (the formation of blood cells). For example, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to increase the quantity of all or subsets of blood cells, such as, for example, erythrocytes, lymphocytes (B or T cells), myeloid cells (e.g., basophils, eosinophils, neutrophils, mast cells, macrophages) and platelets. The ability to decrease the quantity of blood cells or subsets of blood cells may be useful in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of anemias and leukopenias described below. Alternatively, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to decrease the quantity of all or subsets of blood cells, such as, for example, erythrocytes, lymphocytes (B or T cells), myeloid cells (e.g., basophils, eosinophils, neutrophils, mast cells, macrophages) and platelets. The ability to decrease the quantity of blood cells or subsets of blood cells may be useful in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of leukocytoses, such as, for example eosinophilia.

The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate blood dyscrasia.

Anemias are conditions in which the number of red blood cells or amount of hemoglobin (the protein that carries oxygen) in them is below normal. Anemia may be caused by excessive bleeding, decreased red blood cell production, or increased red blood cell destruction (hemolysis). The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating anemias. Anemias that may be treated detect, prevented, diagnosed, prognosticated, treated, and/or ameliorated by the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include iron deficiency anemia, hypochromic anemia, microcytic anemia, chlorosis, hereditary siderob; astic anemia, idiopathic acquired sideroblastic anemia, red cell aplasia, megaloblastic anemia (e.g., pernicious anemia, (vitamin B12 deficiency) and folic acid deficiency anemia), aplastic anemia, hemolytic anemias (e.g., autoimmune helolytic anemia, microangiopathic hemolytic anemia, and paroxysmal nocturnal hemoglobinuria). The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating anemias associated with diseases including but not limited to, anemias associated with systemic lupus erythematosus, cancers, lymphomas, chronic renal disease, and enlarged spleens. The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating anemias arising from drug treatments such as anemias associated with methyldopa, dapsone, and/or sulfadrugs. Additionally, rhe polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating anemias associated with abnormal red blood cell architecture including but not limited to, hereditary spherocytosis, hereditary elliptocytosis, glucose-6-phosphate dehydrogenase deficiency, and sickle cell anemia.

The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating hemoglobin abnormalities, (e.g., those associated with sickle cell anemia, hemoglobin C disease, hemoglobin S—C disease, and hemoglobin E disease). Additionally, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating thalassemias, including, but not limited to major and minor forms of alpha-thalassemia and beta-thalassemia.

In another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating bleeding disorders including, but not limited to, thrombocytopenia (e.g., idiopathic thrombocytopenic purpura, and thrombotic thrombocytopenic purpura), Von Willebrand's disease, hereditary platelet disorders (e.g., storage pool disease such as Chediak-Higashi and Hermansky-Pudlak syndromes, thromboxane A2 dysfunction, thromboasthenia, and Bernard-Soulier syndrome), hemolytic-uremic syndrome, hemophelias such as hemophelia A or Factor VII deficiency and Christmas disease or Factor IX deficiency, Hereditary Hemorhhagic Telangiectsia, also known as Rendu-Osler-Weber syndrome, allergic purpura (Henoch Schonlein purpura) and disseminated intravascular coagulation.

The effect of the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention on the clotting time of blood may be monitored using any of the clotting tests known in the art including, but not limited to, whole blood partial thromboplastin time (PTT), the activated partial thromboplastin time (aPTT), the activated clotting time (ACT), the recalcified activated clotting time, or the Lee-White Clotting time.

Several diseases and a variety of drugs can cause platelet dysfunction. Thus, in a specific embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating acquired platelet dysfunction such as platelet dysfunction accompanying kidney failure, leukemia, multiple myeloma, cirrhosis of the liver, and systemic lupus erythematosus as well as platelet dysfunction associated with drug treatments, including treatment with aspirin, ticlopidine, nonsteroidal anti-inflammatory drugs (used for arthritis, pain, and sprains), and penicillin in high doses.

In another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating diseases and disorders characterized by or associated with increased or decreased numbers of white blood cells. Leukopenia occurs when the number of white blood cells decreases below normal. Leukopenias include, but are not limited to, neutropenia and lymphocytopenia. An increase in the number of white blood cells compared to normal is known as leukocytosis. The body generates increased numbers of white blood cells during infection. Thus, leukocytosis may simply be a normal physiological parameter that reflects infection. Alternatively, leukocytosis may be an indicator of injury or other disease such as cancer. Leokocytoses, include but are not limited to, eosinophilia, and accumulations of macrophages. In specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating leukopenia. In other specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating leukocytosis.

Leukopenia may be a generalized decreased in all types of white blood cells, or may be a specific depletion of particular types of white blood cells. Thus, in specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating decreases in neutrophil numbers, known as neutropenia. Neutropenias that may be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include, but are not limited to, infantile genetic agranulocytosis, familial neutropenia, cyclic neutropenia, neutropenias resulting from or associated with dietary deficiencies (e.g., vitamin B12 deficiency or folic acid deficiency), neutropenias resulting from or associated with drug treatments (e.g., antibiotic regimens such as penicillin treatment, sulfonamide treatment, anticoagulant treatment, anticonvulsant drugs, anti-thyroid drugs, and cancer chemotherapy), and neutropenias resulting from increased neutrophil destruction that may occur in association with some bacterial or viral infections, allergic disorders, autoimmune diseases, conditions in which an individual has an enlarged spleen (e.g., Felty syndrome, malaria and sarcoidosis), and some drug treatment regimens.

The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating lymphocytopenias (decreased numbers of B and/or T lymphocytes), including, but not limited lymphocytopenias resulting from or associated with stress, drug treatments (e.g., drug treatment with corticosteroids, cancer chemotherapies, and/or radiation therapies), AIDS infection and/or other diseases such as, for example, cancer, rheumatoid arthritis, systemic lupus erythematosus, chronic infections, some viral infections and/or hereditary disorders (e.g., DiGeorge syndrome, Wiskott-Aldrich Syndome, severe combined immunodeficiency, ataxia telangiectsia).

The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating diseases and disorders associated with macrophage numbers and/or macrophage function including, but not limited to, Gaucher's disease, Niemann-Pick disease, Letterer-Siwe disease and Hand-Schuller-Christian disease.

In another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating diseases and disorders associated with eosinophil numbers and/or eosinophil function including, but not limited to, idiopathic hypereosinophilic syndrome, eosinophilia-myalgia syndrome, and Hand-S chuller-Christian disease.

In yet another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating leukemias and lymphomas including, but not limited to, acute lymphocytic (lymphpblastic) leukemia (ALL), acute myeloid (myelocytic, myelogenous, myeloblastic, or myelomonocytic) leukemia, chronic lymphocytic leukemia (e.g., B cell leukemias, T cell leukemias, Sezary syndrome, and Hairy cell leukenia), chronic myelocytic (myeloid, myelogenous, or granulocytic) leukemia, Hodgkin's lymphoma, non-hodgkin's lymphoma, Burkitt's lymphoma, and mycosis fungoides.

In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating diseases and disorders of plasma cells including, but not limited to, plasma cell dyscrasias, monoclonal gammaopathies, monoclonal gammopathies of undetermined significance, multiple myeloma, macroglobulinemia, Waldenstrom's macroglobulinemia, cryoglobulinemia, and Raynaud's phenomenon.

In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating myeloproliferative disorders, including but not limited to, polycythemia vera, relative polycythemia, secondary polycythemia, myelofibrosis, acute myelofibrosis, agnogenic myelod metaplasia, thrombocythemia, (including both primary and seconday thrombocythemia) and chronic myelocytic leukemia.

In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as a treatment prior to surgery, to increase blood cell production.

In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to enhance the migration, phagocytosis, superoxide production, antibody dependent cellular cytotoxicity of neutrophils, eosionophils and macrophages.

In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to increase the number of stem cells in circulation prior to stem cells pheresis. In another specific embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to increase the number of stem cells in circulation prior to platelet pheresis.

In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to increase cytokine production.

In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating primary hematopoietic disorders.

Hyperproliferative Disorders

In certain embodiments, polynucleotides or polypeptides, or agonists or antagonists of the present invention can be used to treat or detect hyperproliferative disorders, including neoplasms. Polynucleotides or polypeptides, or agonists or antagonists of the present invention may inhibit the proliferation of the disorder through direct or indirect interactions. Alternatively, Polynucleotides or polypeptides, or agonists or antagonists of the present invention may proliferate other cells which can inhibit the hyperproliferative disorder.

For example, by increasing an immune response, particularly increasing antigenic qualities of the hyperproliferative disorder or by proliferating, differentiating, or mobilizing T-cells, hyperproliferative disorders can be treated. This immune response may be increased by either enhancing an existing immune response, or by initiating a new immune response. Alternatively, decreasing an immune response may also be a method of treating hyperproliferative disorders, such as a chemotherapeutic agent.

Examples of hyperproliferative disorders that can be treated or detected by polynucleotides or polypeptides, or agonists or antagonists of the present invention include, but are not limited to neoplasms located in the: colon, abdomen, bone, breast, digestive system, liver, pancreas, peritoneum, endocrine glands (adrenal, parathyroid, pituitary, testicles, ovary, thymus, thyroid), eye, head and neck, nervous (central and peripheral), lymphatic system, pelvis, skin, soft tissue, spleen, thorax, and urogenital tract.

Similarly, other hyperproliferative disorders can also be treated or detected by polynucleotides or polypeptides, or agonists or antagonists of the present invention. Examples of such hyperproliferative disorders include, but are not limited to: Acute Childhood Lymphoblastic Leukemia, Acute Lymphoblastic Leukemia, Acute Lymphocytic Leukemia, Acute Myeloid Leukemia, Adrenocortical Carcinoma, Adult (Primary) Hepatocellular Cancer, Adult (Primary) Liver Cancer, Adult Acute Lymphocytic Leukemia, Adult Acute Myeloid Leukemia, Adult Hodgkin's Disease, Adult Hodgkin's Lymphoma, Adult Lymphocytic Leukemia, Adult Non-Hodgkin's Lymphoma, Adult Primary Liver Cancer, Adult Soft Tissue Sarcoma, AIDS-Related Lymphoma, AIDS-Related Malignancies, Anal Cancer, Astrocytoma, Bile Duct Cancer, Bladder Cancer, Bone Cancer, Brain Stem Glioma, Brain Tumors, Breast Cancer, Cancer of the Renal Pelvis and Ureter, Central Nervous System (Primary) Lymphoma, Central Nervous System Lymphoma, Cerebellar Astrocytoma, Cerebral Astrocytoma, Cervical Cancer, Childhood (Primary) Hepatocellular Cancer, Childhood (Primary) Liver Cancer, Childhood Acute Lymphoblastic Leukemia, Childhood Acute Myeloid Leukemia, Childhood Brain Stem Glioma, Childhood Cerebellar Astrocytoma, Childhood Cerebral Astrocytoma, Childhood Extracranial Germ Cell Tumors, Childhood Hodgkin's Disease, Childhood Hodgkin's Lymphoma, Childhood Hypothalamic and Visual Pathway Glioma, Childhood Lymphoblastic Leukemia, Childhood Medulloblastoma, Childhood Non-Hodgkin's Lymphoma, Childhood Pineal and Supratentorial Primitive Neuroectodermal Tumors, Childhood Primary Liver Cancer, Childhood Rhabdomyosarcoma, Childhood Soft Tissue Sarcoma, Childhood Visual Pathway and Hypothalamic Glioma, Chronic Lymphocytic Leukemia, Chronic Myelogenous Leukemia, Colon Cancer, Cutaneous T-Cell Lymphoma, Endocrine Pancreas Islet Cell Carcinoma, Endometrial Cancer, Ependymoma, Epithelial Cancer, Esophageal Cancer, Ewing's Sarcoma and Related Tumors, Exocrine Pancreatic Cancer, Extracranial Germ Cell Tumor, Extragonadal Germ Cell Tumor, Extrahepatic Bile Duct Cancer, Eye Cancer, Female Breast Cancer, Gaucher's Disease, Gallbladder Cancer, Gastric Cancer, Gastrointestinal Carcinoid Tumor, Gastrointestinal Tumors, Germ Cell Tumors, Gestational Trophoblastic Tumor, Hairy Cell Leukemia, Head and Neck Cancer, Hepatocellular Cancer, Hodgkin's Disease, Hodgkin's Lymphoma, Hypergammaglobulinemia, Hypopharyngeal Cancer, Intestinal Cancers, Intraocular Melanoma, Islet Cell Carcinoma, Islet Cell Pancreatic Cancer, Kaposi's Sarcoma, Kidney Cancer, Laryngeal Cancer, Lip and Oral Cavity Cancer, Liver Cancer, Lung Cancer, Lymphoproliferative Disorders, Macroglobulinemia, Male Breast Cancer, Malignant Mesothelioma, Malignant Thymoma, Medulloblastoma, Melanoma, Mesothelioma, Metastatic Occult Primary Squamous Neck Cancer, Metastatic Primary Squamous Neck Cancer, Metastatic Squamous Neck Cancer, Multiple Myeloma, Multiple Myeloma/Plasma Cell Neoplasm, Myelodysplastic Syndrome, Myelogenous Leukemia, Myeloid Leukemia, Myeloproliferative Disorders, Nasal Cavity and Paranasal Sinus Cancer, Nasopharyngeal Cancer, Neuroblastoma, Non-Hodgkin's Lymphoma During Pregnancy, Nonmelanoma Skin Cancer, Non-Small Cell Lung Cancer, Occult Primary Metastatic Squamous Neck Cancer, Oropharyngeal Cancer, Osteo-/Malignant Fibrous Sarcoma, Osteosarcoma/Malignant Fibrous Histiocytoma, Osteosarcoma/Malignant Fibrous Histiocytoma of Bone, Ovarian Epithelial Cancer, Ovarian Germ Cell Tumor, Ovarian Low Malignant Potential Tumor, Pancreatic Cancer, Paraproteinemias, Purpura, Parathyroid Cancer, Penile Cancer, Pheochromocytoma, Pituitary Tumor, Plasma Cell Neoplasm/Multiple Myeloma, Primary Central Nervous System Lymphoma, Primary Liver Cancer, Prostate Cancer, Rectal Cancer, Renal Cell Cancer, Renal Pelvis and Ureter Cancer, Retinoblastoma, Rhabdomyosarcoma, Salivary Gland Cancer, Sarcoidosis Sarcomas, Sezary Syndrome, Skin Cancer, Small Cell Lung Cancer, Small Intestine Cancer, Soft Tissue Sarcoma, Squamous Neck Cancer, Stomach Cancer, Supratentorial Primitive Neuroectodermal and Pineal Tumors, T-Cell Lymphoma, Testicular Cancer, Thymoma, Thyroid Cancer, Transitional Cell Cancer of the Renal Pelvis and Ureter, Transitional Renal Pelvis and Ureter Cancer, Trophoblastic Tumors, Ureter and Renal Pelvis Cell Cancer, Urethral Cancer, Uterine Cancer, Uterine Sarcoma, Vaginal Cancer, Visual Pathway and Hypothalamic Glioma, Vulvar Cancer, Waldenstrom's Macroglobulinemia, Wilms' Tumor, and any other hyperproliferative disease, besides neoplasia, located in an organ system listed above.

In another preferred embodiment, polynucleotides or polypeptides, or agonists or antagonists of the present invention are used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate premalignant conditions and to prevent progression to a neoplastic or malignant state, including but not limited to those disorders described above. Such uses are indicated in conditions known or suspected of preceding progression to neoplasia or cancer, in particular, where non-neoplastic cell growth consisting of hyperplasia, metaplasia, or most particularly, dysplasia has occurred (for review of such abnormal growth conditions, see Robbins and Angell, 1976, Basic Pathology, 2d Ed., W. B. Saunders Co., Philadelphia, pp. 68-79.)

Hyperplasia is a form of controlled cell proliferation, involving an increase in cell number in a tissue or organ, without significant alteration in structure or function. Hyperplastic disorders which can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, angiofollicular mediastinal lymph node hyperplasia, angiolymphoid hyperplasia with eosinophilia, atypical melanocytic hyperplasia, basal cell hyperplasia, benign giant lymph node hyperplasia, cementum hyperplasia, congenital adrenal hyperplasia, congenital sebaceous hyperplasia, cystic hyperplasia, cystic hyperplasia of the breast, denture hyperplasia, ductal hyperplasia, endometrial hyperplasia, fibromuscular hyperplasia, focal epithelial hyperplasia, gingival hyperplasia, inflammatory fibrous hyperplasia, inflammatory papillary hyperplasia, intravascular papillary endothelial hyperplasia, nodular hyperplasia of prostate, nodular regenerative hyperplasia, pseudoepitheliomatous hyperplasia, senile sebaceous hyperplasia, and verrucous hyperplasia.

Metaplasia is a form of controlled cell growth in which one type of adult or fully differentiated cell substitutes for another type of adult cell. Metaplastic disorders which can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, agnogenic myeloid metaplasia, apocrine metaplasia, atypical metaplasia, autoparenchymatous metaplasia, connective tissue metaplasia, epithelial metaplasia, intestinal metaplasia, metaplastic anemia, metaplastic ossification, metaplastic polyps, myeloid metaplasia, primary myeloid metaplasia, secondary myeloid metaplasia, squamous metaplasia, squamous metaplasia of amnion, and symptomatic myeloid metaplasia.

Dysplasia is frequently a forerunner of cancer, and is found mainly in the epithelia; it is the most disorderly form of non-neoplastic cell growth, involving a loss in individual cell uniformity and in the architectural orientation of cells. Dysplastic cells often have abnormally large, deeply stained nuclei, and exhibit pleomorphism. Dysplasia characteristically occurs where there exists chronic irritation or inflammation. Dysplastic disorders which can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, anhidrotic ectodermal dysplasia, anterofacial dysplasia, asphyxiating thoracic dysplasia, atriodigital dysplasia, bronchopulmonary dysplasia, cerebral dysplasia, cervical dysplasia, chondroectodermal dysplasia, cleidocranial dysplasia, congenital ectodermal dysplasia, craniodiaphysial dysplasia, craniocarpotarsal dysplasia, craniometaphysial dysplasia, dentin dysplasia, diaphysial dysplasia, ectodermal dysplasia, enamel dysplasia, encephalo-ophthalmic dysplasia, dysplasia epiphysialis hemimelia, dysplasia epiphysialis multiplex, dysplasia epiphysialis punctata, epithelial dysplasia, faciodigitogenital dysplasia, familial fibrous dysplasia of jaws, familial white folded dysplasia, fibromuscular dysplasia, fibrous dysplasia of bone, florid osseous dysplasia, hereditary renal-retinal dysplasia, hidrotic ectodermal dysplasia, hypohidrotic ectodermal dysplasia, lymphopenic thymic dysplasia, mammary dysplasia, mandibulofacial dysplasia, metaphysial dysplasia, Mondini dysplasia, monostotic fibrous dysplasia, mucoepithelial dysplasia, multiple epiphysial dysplasia, oculoauriculovertebral dysplasia, oculodentodigital dysplasia, oculovertebral dysplasia, odontogenic dysplasia, ophthalmomandibulomelic dysplasia, periapical cemental dysplasia, polyostotic fibrous dysplasia, pseudoachondroplastic spondyloepiphysial dysplasia, retinal dysplasia, septo-optic dysplasia, spondyloepiphysial dysplasia, and ventriculoradial dysplasia.

Additional pre-neoplastic disorders which can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, benign dysproliferative disorders (e.g., benign tumors, fibrocystic conditions, tissue hypertrophy, intestinal polyps, colon polyps, and esophageal dysplasia), leukoplakia, keratoses, Bowen's disease, Farmer's Skin, solar cheilitis, and solar keratosis.

In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to diagnose and/or prognosticate disorders associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1B.2, column 5 (Tissue Distribution Library Code).

In another embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention conjugated to a toxin or a radioactive isotope, as described herein, may be used to treat cancers and neoplasms, including, but not limited to those described herein. In a further preferred embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention conjugated to a toxin or a radioactive isotope, as described herein, may be used to treat acute myelogenous leukemia.

Additionally, polynucleotides, polypeptides, and/or agonists or antagonists of the invention may affect apoptosis, and therefore, would be useful in treating a number of diseases associated with increased cell survival or the inhibition of apoptosis. For example, diseases associated with increased cell survival or the inhibition of apoptosis that could be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include cancers (such as follicular lymphomas, carcinomas with p53 mutations, and hormone-dependent tumors, including, but not limited to colon cancer, cardiac tumors, pancreatic cancer, melanoma, retinoblastoma, glioblastoma, lung cancer, intestinal cancer, testicular cancer, stomach cancer, neuroblastoma, myxoma, myoma, lymphoma, endothelioma, osteoblastoma, osteoclastoma, osteosarcoma, chondrosarcoma, adenoma, breast cancer, prostate cancer, Kaposi's sarcoma and ovarian cancer); autoimmune disorders such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) and viral infections (such as herpes viruses, pox viruses and adenoviruses), inflammation, graft v. host disease, acute graft rejection, and chronic graft rejection.

In preferred embodiments, polynucleotides, polypeptides, and/or agonists or antagonists of the invention are used to inhibit growth, progression, and/or metastasis of cancers, in particular those listed above.

Additional diseases or conditions associated with increased cell survival that could be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include, but are not limited to, progression, and/or metastases of malignancies and related disorders such as leukemia (including acute leukemias (e.g., acute lymphocytic leukemia, acute myelocytic leukemia (including myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia)) and chronic leukemias (e.g., chronic myelocytic (granulocytic) leukemia and chronic lymphocytic leukemia)), polycythemia vera, lymphomas (e.g., Hodgkin's disease and non-Hodgkin's disease), multiple myeloma, Waldenstrom's macroglobulinemia, heavy chain disease, and solid tumors including, but not limited to, sarcomas and carcinomas such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm's tumor, cervical cancer, testicular tumor, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, emangioblastoma, acoustic neuroma, oligodendroglioma, menangioma, melanoma, neuroblastoma, and retinoblastoma.

Diseases associated with increased apoptosis that could be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include AIDS; neurodegenerative disorders (such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, retinitis pigmentosa, cerebellar degeneration and brain tumor or prior associated disease); autoimmune disorders (such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) myelodysplastic syndromes (such as aplastic anemia), graft v. host disease, ischemic injury (such as that caused by myocardial infarction, stroke and reperfusion injury), liver injury (e.g., hepatitis related liver injury, ischemia/reperfusion injury, cholestosis (bile duct injury) and liver cancer); toxin-induced liver disease (such as that caused by alcohol), septic shock, cachexia and anorexia.

Hyperproliferative diseases and/or disorders that could be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include, but are not limited to, neoplasms located in the liver, abdomen, bone, breast, digestive system, pancreas, peritoneum, endocrine glands (adrenal, parathyroid, pituitary, testicles, ovary, thymus, thyroid), eye, head and neck, nervous system (central and peripheral), lymphatic system, pelvis, skin, soft tissue, spleen, thorax, and urogenital tract.

Similarly, other hyperproliferative disorders can also be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention. Examples of such hyperproliferative disorders include, but are not limited to: hypergammaglobulinemia, lymphoproliferative disorders, paraproteinemias, purpura, sarcoidosis, Sezary Syndrome, Waldenstron's macroglobulinemia, Gaucher's Disease, histiocytosis, and any other hyperproliferative disease, besides neoplasia, located in an organ system listed above.

Another preferred embodiment utilizes polynucleotides of the present invention to inhibit aberrant cellular division, by gene therapy using the present invention, and/or protein fusions or fragments thereof.

Thus, the present invention provides a method for treating cell proliferative disorders by inserting into an abnormally proliferating cell a polynucleotide of the present invention, wherein said polynucleotide represses said expression.

Another embodiment of the present invention provides a method of treating cell-proliferative disorders in individuals comprising administration of one or more active gene copies of the present invention to an abnormally proliferating cell or cells. In a preferred embodiment, polynucleotides of the present invention is a DNA construct comprising a recombinant expression vector effective in expressing a DNA sequence encoding said polynucleotides. In another preferred embodiment of the present invention, the DNA construct encoding the polynucleotides of the present invention is inserted into cells to be treated utilizing a retrovirus, or more preferably an adenoviral vector (See G J. Nabel, et. al., PNAS 1999 96: 324-326, which is hereby incorporated by reference). In a most preferred embodiment, the viral vector is defective and will not transform non-proliferating cells, only proliferating cells. Moreover, in a preferred embodiment, the polynucleotides of the present invention inserted into proliferating cells either alone, or in combination with or fused to other polynucleotides, can then be modulated via an external stimulus (i.e. magnetic, specific small molecule, chemical, or drug administration, etc.), which acts upon the promoter upstream of said polynucleotides to induce expression of the encoded protein product. As such the beneficial therapeutic affect of the present invention may be expressly modulated (i.e. to increase, decrease, or inhibit expression of the present invention) based upon said external stimulus.

Polynucleotides of the present invention may be useful in repressing expression of oncogenic genes or antigens. By “repressing expression of the oncogenic genes” is intended the suppression of the transcription of the gene, the degradation of the gene transcript (pre-message RNA), the inhibition of splicing, the destruction of the messenger RNA, the prevention of the post-translational modifications of the protein, the destruction of the protein, or the inhibition of the normal function of the protein.

For local administration to abnormally proliferating cells, polynucleotides of the present invention may be administered by any method known to those of skill in the art including, but not limited to transfection, electroporation, microinjection of cells, or in vehicles such as liposomes, LIPOFECTIN™, or as naked polynucleotides, or any other method described throughout the specification. The polynucleotide of the present invention may be delivered by known gene delivery systems such as, but not limited to, retroviral vectors (Gilboa, J. Virology 44:845 (1982); Hocke, Nature 320:275 (1986); Wilson, et al., Proc. Natl. Acad. Sci. U.S.A. 85:3014), vaccinia virus system (Chakrabarty et al., Mol. Cell. Biol. 5:3403 (1985) or other efficient DNA delivery systems (Yates et al., Nature 313:812 (1985)) known to those skilled in the art. These references are exemplary only and are hereby incorporated by reference. In order to specifically deliver or transfect cells which are abnormally proliferating and spare non-dividing cells, it is preferable to utilize a retrovirus, or adenoviral (as described in the art and elsewhere herein) delivery system known to those of skill in the art. Since host DNA replication is required for retroviral DNA to integrate and the retrovirus will be unable to self replicate due to the lack of the retrovirus genes needed for its life cycle. Utilizing such a retroviral delivery system for polynucleotides of the present invention will target said gene and constructs to abnormally proliferating cells and will spare the non-dividing normal cells.

The polynucleotides of the present invention may be delivered directly to cell proliferative disorder/disease sites in internal organs, body cavities and the like by use of imaging devices used to guide an injecting needle directly to the disease site. The polynucleotides of the present invention may also be administered to disease sites at the time of surgical intervention.

By “cell proliferative disease” is meant any human or animal disease or disorder, affecting any one or any combination of organs, cavities, or body parts, which is characterized by single or multiple local abnormal proliferations of cells, groups of cells, or tissues, whether benign or malignant.

Any amount of the polynucleotides of the present invention may be administered as long as it has a biologically inhibiting effect on the proliferation of the treated cells. Moreover, it is possible to administer more than one of the polynucleotide of the present invention simultaneously to the same site. By “biologically inhibiting” is meant partial or total growth inhibition as well as decreases in the rate of proliferation or growth of the cells. The biologically inhibitory dose may be determined by assessing the effects of the polynucleotides of the present invention on target malignant or abnormally proliferating cell growth in tissue culture, tumor growth in animals and cell cultures, or any other method known to one of ordinary skill in the art.

The present invention is further directed to antibody-based therapies which involve administering of anti-polypeptides and anti-polynucleotide antibodies to a mammalian, preferably human, patient for treating one or more of the described disorders. Methods for producing anti-polypeptides and anti-polynucleotide antibodies polyclonal and monoclonal antibodies are described in detail elsewhere herein. Such antibodies may be provided in pharmaceutically acceptable compositions as known in the art or as described herein.

A summary of the ways in which the antibodies of the present invention may be used therapeutically includes binding polynucleotides or polypeptides of the present invention locally or systemically in the body or by direct cytotoxicity of the antibody, e.g. as mediated by complement (CDC) or by effector cells (ADCC). Some of these approaches are described in more detail below. Armed with the teachings provided herein, one of ordinary skill in the art will know how to use the antibodies of the present invention for diagnosis, prognosis, monitoring, or therapeutic purposes without undue experimentation.

In particular, the antibodies, fragments and derivatives of the present invention are useful for treating a subject having or developing cell proliferative and/or differentiation disorders as described herein. Such treatment comprises administering a single or multiple doses of the antibody, or a fragment, derivative, or a conjugate thereof.

The antibodies of this invention may be advantageously utilized in combination with other monoclonal or chimeric antibodies, or with lymphokines or hematopoietic growth factors, for example., which serve to increase the number or activity of effector cells which interact with the antibodies.

It is preferred to use high affinity and/or potent in vivo inhibiting and/or neutralizing antibodies against polypeptides or polynucleotides of the present invention, fragments or regions thereof, for both immunoassays directed to and therapy of disorders related to polynucleotides or polypeptides, including fragments thereof, of the present invention. Such antibodies, fragments, or regions, will preferably have an affinity for polynucleotides or polypeptides, including fragments thereof. Preferred binding affinities include those with a dissociation constant or Kd less than 5×10⁶M, 10⁻⁶M, 5×10⁷M, 10⁻⁷M, 5×10⁸M, 10⁻⁸M, 5×10⁹M, 10⁻⁹M, 5×10⁻¹⁰M, 10⁻¹⁰M, 5×10⁻¹¹M, 10⁻¹¹M, 5×10⁻¹²M, 10⁻¹²M, 5×10⁻¹³M, 10⁻¹³M, 5×10⁻¹⁴M, 10⁻¹⁴M, 5×10⁻¹⁵M, and 10⁻¹⁵M.

Moreover, polypeptides of the present invention are useful in inhibiting the angiogenesis of proliferative cells or tissues, either alone, as a protein fusion, or in combination with other polypeptides directly or indirectly, as described elsewhere herein. In a most preferred embodiment, said anti-angiogenesis effect may be achieved indirectly, for example, through the inhibition of hematopoietic, tumor-specific cells, such as tumor-associated macrophages (See Joseph I B, et al. J Natl Cancer Inst, 90(21):1648-53 (1998), which is hereby incorporated by reference). Antibodies directed to polypeptides or polynucleotides of the present invention may also result in inhibition of angiogenesis directly, or indirectly (See Witte L, et al., Cancer Metastasis Rev. 17(2):155-61 (1998), which is hereby incorporated by reference)).

Polypeptides, including protein fusions, of the present invention, or fragments thereof may be useful in inhibiting proliferative cells or tissues through the induction of apoptosis. Said polypeptides may act either directly, or indirectly to induce apoptosis of proliferative cells and tissues, for example in the activation of a death-domain receptor, such as tumor necrosis factor (TNF) receptor-1, CD95 (Fas/APO-1), TNF-receptor-related apoptosis-mediated protein (TRAMP) and TNF-related apoptosis-inducing ligand (TRAIL) receptor-1 and -2 (See Schulze-Osthoff K, et. al., Eur J Biochem 254(3):439-59 (1998), which is hereby incorporated by reference). Moreover, in another preferred embodiment of the present invention, said polypeptides may induce apoptosis through other mechanisms, such as in the activation of other proteins which will activate apoptosis, or through stimulating the expression of said proteins, either alone or in combination with small molecule drugs or adjuviants, such as apoptonin, galectins, thioredoxins, anti-inflammatory proteins (See for example, Mutat Res 400(1-2):447-55 (1998), Med. Hypotheses.50(5):423-33 (1998), Chem Biol Interact. April 24; 111-112:23-34 (1998), J Mol. Med. 76(6):402-12 (1998), Int J Tissue React;20(1):3-15 (1998), which are all hereby incorporated by reference).

Polypeptides, including protein fusions to, or fragments thereof, of the present invention are useful in inhibiting the metastasis of proliferative cells or tissues Inhibition may occur as a direct result of administering polypeptides, or antibodies directed to said polypeptides as described elsewhere herein, or indirectly, such as activating the expression of proteins known to inhibit metastasis, for example alpha 4 integrins, (See, e.g., Curr Top Microbiol Immunol 1998; 231:125-41, which is hereby incorporated by reference). Such therapeutic affects of the present invention may be achieved either alone, or in combination with small molecule drugs or adjuvants.

In another embodiment, the invention provides a method of delivering compositions containing the polypeptides of the invention (e.g., compositions containing polypeptides or polypeptide antibodies associated with heterologous polypeptides, heterologous nucleic acids, toxins, or prodrugs) to targeted cells expressing the polypeptide of the present invention. Polypeptides or polypeptide antibodies of the invention may be associated with heterologous polypeptides, heterologous nucleic acids, toxins, or prodrugs via hydrophobic, hydrophilic, ionic and/or covalent interactions.

Polypeptides, protein fusions to, or fragments thereof, of the present invention are useful in enhancing the immunogenicity and/or antigenicity of proliferating cells or tissues, either directly, such as would occur if the polypeptides of the present invention ‘vaccinated’ the immune response to respond to proliferative antigens and immunogens, or indirectly, such as in activating the expression of proteins known to enhance the immune response (e.g. chemokines), to said antigens and immunogens.

Renal Disorders

Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention, may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate disorders of the renal system. Renal disorders which can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with compositions of the invention include, but are not limited to, kidney failure, nephritis, blood vessel disorders of kidney, metabolic and congenital kidney disorders, urinary disorders of the kidney, autoimmune disorders, sclerosis and necrosis, electrolyte imbalance, and kidney cancers.

Kidney diseases which can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with compositions of the invention include, but are not limited to, acute kidney failure, chronic kidney failure, atheroembolic renal failure, end-stage renal disease, inflammatory diseases of the kidney (e.g., acute glomerulonephritis, postinfectious glomerulonephritis, rapidly progressive glomerulonephritis, nephrotic syndrome, membranous glomerulonephritis, familial nephrotic syndrome, membranoproliferative glomerulonephritis I and II, mesangial proliferative glomerulonephritis, chronic glomerulonephritis, acute tubulointerstitial nephritis, chronic tubulointerstitial nephritis, acute post-streptococcal glomerulonephritis (PSGN), pyelonephritis, lupus nephritis, chronic nephritis, interstitial nephritis, and post-streptococcal glomerulonephritis), blood vessel disorders of the kidneys (e.g., kidney infarction, atheroembolic kidney disease, cortical necrosis, malignant nephrosclerosis, renal vein thrombosis, renal underperfusion, renal retinopathy, renal ischemia-reperfusion, renal artery embolism, and renal artery stenosis), and kidney disorders resulting form urinary tract disease (e.g., pyelonephritis, hydronephrosis, urolithiasis (renal lithiasis, nephrolithiasis), reflux nephropathy, urinary tract infections, urinary retention, and acute or chronic unilateral obstructive uropathy.)

In addition, compositions of the invention can be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate metabolic and congenital disorders of the kidney (e.g., uremia, renal amyloidosis, renal osteodystrophy, renal tubular acidosis, renal glycosuria, nephrogenic diabetes insipidus, cystinuria, Fanconi's syndrome, renal fibrocystic osteosis (renal rickets), Hartnup disease, Bartter's syndrome, Liddle's syndrome, polycystic kidney disease, medullary cystic disease, medullary sponge kidney, Alport's syndrome, nail-patella syndrome, congenital nephrotic syndrome, CRUSH syndrome, horseshoe kidney, diabetic nephropathy, nephrogenic diabetes insipidus, analgesic nephropathy, kidney stones, and membranous nephropathy), and autoimmune disorders of the kidney (e.g., systemic lupus erythematosus (SLE), Goodpasture syndrome, IgA nephropathy, and IgM mesangial proliferative glomerulonephritis).

Compositions of the invention can also be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate sclerotic or necrotic disorders of the kidney (e.g., glomerulosclerosis, diabetic nephropathy, focal segmental glomerulosclerosis (FSGS), necrotizing glomerulonephritis, and renal papillary necrosis), cancers of the kidney (e.g., nephroma, hypernephroma, nephroblastoma, renal cell cancer, transitional cell cancer, renal adenocarcinoma, squamous cell cancer, and Wilm's tumor), and electrolyte imbalances (e.g., nephrocalcinosis, pyuria, edema, hydronephritis, proteinuria, hyponatremia, hypernatremia, hypokalemia, hyperkalemia, hypocalcemia, hypercalcemia, hypophosphatemia, and hyperphosphatemia).

Polypeptides may be administered using any method known in the art, including, but not limited to, direct needle injection at the delivery site, intravenous injection, topical administration, catheter infusion, biolistic injectors, particle accelerators, gelfoam sponge depots, other commercially available depot materials, osmotic pumps, oral or suppositorial solid pharmaceutical formulations, decanting or topical applications during surgery, aerosol delivery. Such methods are known in the art. Polypeptides may be administered as part of a Therapeutic, described in more detail below. Methods of delivering polynucleotides are described in more detail herein.

Cardiovascular Disorders

Polynucleotides or polypeptides, or agonists or antagonists of the present invention, may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate cardiovascular diseases and disorders, including, but not limited to, peripheral artery disease, such as limb ischemia.

Cardiovascular disorders include, but are not limited to, cardiovascular abnormalities, such as arterio-arterial fistula, arteriovenous fistula, cerebral arteriovenous malformations, congenital heart defects, pulmonary atresia, and Scimitar Syndrome. Congenital heart defects include, but are not limited to, aortic coarctation, cor triatriatum, coronary vessel anomalies, crisscross heart, dextrocardia, patent ductus arteriosus, Ebstein's anomaly, Eisenmenger complex, hypoplastic left heart syndrome, levocardia, tetralogy of fallot, transposition of great vessels, double outlet right ventricle, tricuspid atresia, persistent truncus arteriosus, and heart septal defects, such as aortopulmonary septal defect, endocardial cushion defects, Lutembacher's Syndrome, trilogy of Fallot, ventricular heart septal defects.

Cardiovascular disorders also include, but are not limited to, heart disease, such as arrhythmias, carcinoid heart disease, high cardiac output, low cardiac output, cardiac tamponade, endocarditis (including bacterial), heart aneurysm, cardiac arrest, congestive heart failure, congestive cardiomyopathy, paroxysmal dyspnea, cardiac edema, heart hypertrophy, congestive cardiomyopathy, left ventricular hypertrophy, right ventricular hypertrophy, post-infarction heart rupture, ventricular septal rupture, heart valve diseases, myocardial diseases, myocardial ischemia, pericardial effusion, pericarditis (including constrictive and tuberculous), pneumopericardium, postpericardiotomy syndrome, pulmonary heart disease, rheumatic heart disease, ventricular dysfunction, hyperemia, cardiovascular pregnancy complications, Scimitar Syndrome, cardiovascular syphilis, and cardiovascular tuberculosis.

Arrhythmias include, but are not limited to, sinus arrhythmia, atrial fibrillation, atrial flutter, bradycardia, extrasystole, Adams-Stokes Syndrome, bundle-branch block, sinoatrial block, long QT syndrome, parasystole, Lown-Ganong-Levine Syndrome, Mahaim-type pre-excitation syndrome, Wolff-Parkinson-White syndrome, sick sinus syndrome, tachycardias, and ventricular fibrillation. Tachycardias include paroxysmal tachycardia, supraventricular tachycardia, accelerated idioventricular rhythm, atrioventricular nodal reentry tachycardia, ectopic atrial tachycardia, ectopic junctional tachycardia, sinoatrial nodal reentry tachycardia, sinus tachycardia, Torsades de Pointes, and ventricular tachycardia.

Heart valve diseases include, but are not limited to, aortic valve insufficiency, aortic valve stenosis, hear murmurs, aortic valve prolapse, mitral valve prolapse, tricuspid valve prolapse, mitral valve insufficiency, mitral valve stenosis, pulmonary atresia, pulmonary valve insufficiency, pulmonary valve stenosis, tricuspid atresia, tricuspid valve insufficiency, and tricuspid valve stenosis.

Myocardial diseases include, but are not limited to, alcoholic cardiomyopathy, congestive cardiomyopathy, hypertrophic cardiomyopathy, aortic subvalvular stenosis, pulmonary subvalvular stenosis, restrictive cardiomyopathy, Chagas cardiomyopathy, endocardial fibroelastosis, endomyocardial fibrosis, Kearns Syndrome, myocardial reperfusion injury, and myocarditis.

Myocardial ischemias include, but are not limited to, coronary disease, such as angina pectoris, coronary aneurysm, coronary arteriosclerosis, coronary thrombosis, coronary vasospasm, myocardial infarction and myocardial stunning.

Cardiovascular diseases also include vascular diseases such as aneurysms, angiodysplasia, angiomatosis, bacillary angiomatosis, Hippel-Lindau Disease, Klippel-Trenaunay-Weber Syndrome, Sturge-Weber Syndrome, angioneurotic edema, aortic diseases, Takayasu's Arteritis, aortitis, Leriche's Syndrome, arterial occlusive diseases, arteritis, enarteritis, polyarteritis nodosa, cerebrovascular disorders, diabetic angiopathies, diabetic retinopathy, embolisms, thrombosis, erythromelalgia, hemorrhoids, hepatic veno-occlusive disease, hypertension, hypotension, ischemia, peripheral vascular diseases, phlebitis, pulmonary veno-occlusive disease, Raynaud's disease, CREST syndrome, retinal vein occlusion, Scimitar syndrome, superior vena cava syndrome, telangiectasia, atacia telangiectasia, hereditary hemorrhagic telangiectasia, varicocele, varicose veins, varicose ulcer, vasculitis, and venous insufficiency.

Aneurysms include, but are not limited to, dissecting aneurysms, false aneurysms, infected aneurysms, ruptured aneurysms, aortic aneurysms, cerebral aneurysms, coronary aneurysms, heart aneurysms, and iliac aneurysms.

Arterial occlusive diseases include, but are not limited to, arteriosclerosis, intermittent claudication, carotid stenosis, fibromuscular dysplasias, mesenteric vascular occlusion, Moyamoya disease, renal artery obstruction, retinal artery occlusion, and thromboangiitis obliterans.

Cerebrovascular disorders include, but are not limited to, carotid artery diseases, cerebral amyloid angiopathy, cerebral aneurysm, cerebral anoxia, cerebral arteriosclerosis, cerebral arteriovenous malformation, cerebral artery diseases, cerebral embolism and thrombosis, carotid artery thrombosis, sinus thrombosis, Wallenberg's syndrome, cerebral hemorrhage, epidural hematoma, subdural hematoma, subaraxhnoid hemorrhage, cerebral infarction, cerebral ischemia (including transient), subclavian steal syndrome, periventricular leukomalacia, vascular headache, cluster headache, migraine, and vertebrobasilar insufficiency.

Embolisms include, but are not limited to, air embolisms, amniotic fluid embolisms, cholesterol embolisms, blue toe syndrome, fat embolisms, pulmonary embolisms, and thromoboembolisms. Thrombosis include, but are not limited to, coronary thrombosis, hepatic vein thrombosis, retinal vein occlusion, carotid artery thrombosis, sinus thrombosis, Wallenberg's syndrome, and thrombophlebitis.

Ischemic disorders include, but are not limited to, cerebral ischemia, ischemic colitis, compartment syndromes, anterior compartment syndrome, myocardial ischemia, reperfusion injuries, and peripheral limb ischemia. Vasculitis includes, but is not limited to, aortitis, arteritis, Behcet's Syndrome, Churg-Strauss Syndrome, mucocutaneous lymph node syndrome, thromboangiitis obliterans, hypersensitivity vasculitis, Schoenlein-Henoch purpura, allergic cutaneous vasculitis, and Wegener's granulomatosis.

Polypeptides may be administered using any method known in the art, including, but not limited to, direct needle injection at the delivery site, intravenous injection, topical administration, catheter infusion, biolistic injectors, particle accelerators, gelfoam sponge depots, other commercially available depot materials, osmotic pumps, oral or suppositorial solid pharmaceutical formulations, decanting or topical applications during surgery, aerosol delivery. Such methods are known in the art. Polypeptides may be administered as part of a Therapeutic, described in more detail below. Methods of delivering polynucleotides are described in more detail herein.

Respiratory Disorders

Polynucleotides or polypeptides, or agonists or antagonists of the present invention may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate diseases and/or disorders of the respiratory system.

Diseases and disorders of the respiratory system include, but are not limited to, nasal vestibulitis, nonallergic rhinitis (e.g., acute rhinitis, chronic rhinitis, atrophic rhinitis, vasomotor rhinitis), nasal polyps, and sinusitis, juvenile angiofibromas, cancer of the nose and juvenile papillomas, vocal cord polyps, nodules (singer's nodules), contact ulcers, vocal cord paralysis, laryngoceles, pharyngitis (e.g., viral and bacterial), tonsillitis, tonsillar cellulitis, parapharyngeal abscess, laryngitis, laryngoceles, and throat cancers (e.g., cancer of the nasopharynx, tonsil cancer, larynx cancer), lung cancer (e.g., squamous cell carcinoma, small cell (oat cell) carcinoma, large cell carcinoma, and adenocarcinoma), allergic disorders (eosinophilic pneumonia, hypersensitivity pneumonitis (e.g., extrinsic allergic alveolitis, allergic interstitial pneumonitis, organic dust pneumoconiosis, allergic bronchopulmonary aspergillosis, asthma, Wegener's granulomatosis (granulomatous vasculitis), Goodpasture's syndrome)), pneumonia (e.g., bacterial pneumonia (e.g., Streptococcus pneumoniae (pneumoncoccal pneumonia), Staphylococcus aureus (staphylococcal pneumonia), Gram-negative bacterial pneumonia (caused by, e.g., Klebsiella and Pseudomas spp.), Mycoplasma pneumoniae pneumonia, Hemophilus influenzae pneumonia, Legionella pneumophila (Legionnaires' disease), and Chlamydia psittaci (Psittacosis)), and viral pneumonia (e.g., influenza, chickenpox (varicella).

Additional diseases and disorders of the respiratory system include, but are not limited to bronchiolitis, polio (poliomyelitis), croup, respiratory syncytial viral infection, mumps, erythema infectiosum (fifth disease), roseola infantum, progressive rubella panencephalitis, german measles, and subacute sclerosing panencephalitis), fungal pneumonia (e.g., Histoplasmosis, Coccidioidomycosis, Blastomycosis, fungal infections in people with severely suppressed immune systems (e.g., cryptococcosis, caused by Cryptococcus neoformans; aspergillosis, caused by Aspergillus spp.; candidiasis, caused by Candida; and mucormycosis)), Pneumocystis carinii (pneumocystis pneumonia), atypical pneumonias (e.g., Mycoplasma and Chlamydia spp.), opportunistic infection pneumonia, nosocomial pneumonia, chemical pneumonitis, and aspiration pneumonia, pleural disorders (e.g., pleurisy, pleural effusion, and pneumothorax (e.g., simple spontaneous pneumothorax, complicated spontaneous pneumothorax, tension pneumothorax)), obstructive airway diseases (e.g., asthma, chronic obstructive pulmonary disease (COPD), emphysema, chronic or acute bronchitis), occupational lung diseases (e.g., silicosis, black lung (coal workers' pneumoconiosis), asbestosis, berylliosis, occupational asthsma, byssinosis, and benign pneumoconioses), Infiltrative Lung Disease (e.g., pulmonary fibrosis (e.g., fibrosing alveolitis, usual interstitial pneumonia), idiopathic pulmonary fibrosis, desquamative interstitial pneumonia, lymphoid interstitial pneumonia, histiocytosis X (e.g., Letterer-Siwe disease, Hand-Schüller-Christian disease, eosinophilic granuloma), idiopathic pulmonary hemosiderosis, sarcoidosis and pulmonary alveolar proteinosis), Acute respiratory distress syndrome (also called, e.g., adult respiratory distress syndrome), edema, pulmonary embolism, bronchitis (e.g., viral, bacterial), bronchiectasis, atelectasis, lung abscess (caused by, e.g., Staphylococcus aureus or Legionella pneumophila), and cystic fibrosis.

Anti-Angiogenesis Activity

The naturally occurring balance between endogenous stimulators and inhibitors of angiogenesis is one in which inhibitory influences predominate. Rastinejad et al., Cell 56:345-355 (1989). In those rare instances in which neovascularization occurs under normal physiological conditions, such as wound healing, organ regeneration, embryonic development, and female reproductive processes, angiogenesis is stringently regulated and spatially and temporally delimited. Under conditions of pathological angiogenesis such as that characterizing solid tumor growth, these regulatory controls fail. Unregulated angiogenesis becomes pathologic and sustains progression of many neoplastic and non-neoplastic diseases. A number of serious diseases are dominated by abnormal neovascularization including solid tumor growth and metastases, arthritis, some types of eye disorders, and psoriasis. See, e.g., reviews by Moses et al., Biotech. 9:630-634 (1991); Folkman et al., N. Engl. J. Med., 333:1757-1763 (1995); Auerbach et al., J. Microvasc. Res. 29:401-411 (1985); Folkman, Advances in Cancer Research, eds. Klein and Weinhouse, Academic Press, New York, pp. 175-203 (1985); Patz, Am. J. Opthalmol. 94:715-743 (1982); and Folkman et al., Science 221:719-725 (1983). In a number of pathological conditions, the process of angiogenesis contributes to the disease state. For example, significant data have accumulated which suggest that the growth of solid tumors is dependent on angiogenesis. Folkman and Klagsbrun, Science 235:442-447 (1987).

The present invention provides for treatment of diseases or disorders associated with neovascularization by administration of the polynucleotides and/or polypeptides of the invention, as well as agonists or antagonists of the present invention. Malignant and metastatic conditions which can be treated with the polynucleotides and polypeptides, or agonists or antagonists of the invention include, but are not limited to, malignancies, solid tumors, and cancers described herein and otherwise known in the art (for a review of such disorders, see Fishman et al., Medicine, 2d Ed., J. B. Lippincott Co., Philadelphia (1985)). Thus, the present invention provides a method of treating an angiogenesis-related disease and/or disorder, comprising administering to an individual in need thereof a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist of the invention. For example, polynucleotides, polypeptides, antagonists and/or agonists may be utilized in a variety of additional methods in order to therapeutically treat a cancer or tumor. Cancers which may be treated with polynucleotides, polypeptides, antagonists and/or agonists include, but are not limited to solid tumors, including prostate, lung, breast, ovarian, stomach, pancreas, larynx, esophagus, testes, liver, parotid, biliary tract, colon, rectum, cervix, uterus, endometrium, kidney, bladder, thyroid cancer; primary tumors and metastases; melanomas; glioblastoma; Kaposi's sarcoma; leiomyosarcoma; non-small cell lung cancer; colorectal cancer; advanced malignancies; and blood born tumors such as leukemias. For example, polynucleotides, polypeptides, antagonists and/or agonists may be delivered topically, in order to treat cancers such as skin cancer, head and neck tumors, breast tumors, and Kaposi's sarcoma.

Within yet other aspects, polynucleotides, polypeptides, antagonists and/or agonists may be utilized to treat superficial forms of bladder cancer by, for example, intravesical administration. Polynucleotides, polypeptides, antagonists and/or agonists may be delivered directly into the tumor, or near the tumor site, via injection or a catheter. Of course, as the artisan of ordinary skill will appreciate, the appropriate mode of administration will vary according to the cancer to be treated. Other modes of delivery are discussed herein.

Polynucleotides, polypeptides, antagonists and/or agonists may be useful in treating other disorders, besides cancers, which involve angiogenesis. These disorders include, but are not limited to: benign tumors, for example hemangiomas, acoustic neuromas, neurofibromas, trachomas, and pyogenic granulomas; artheroscleric plaques; ocular angiogenic diseases, for example, diabetic retinopathy, retinopathy of prematurity, macular degeneration, corneal graft rejection, neovascular glaucoma, retrolental fibroplasia, rubeosis, retinoblastoma, uvietis and Pterygia (abnormal blood vessel growth) of the eye; rheumatoid arthritis; psoriasis; delayed wound healing; endometriosis; vasculogenesis; granulations; hypertrophic scars (keloids); nonunion fractures; scleroderma; trachoma; vascular adhesions; myocardial angiogenesis; coronary collaterals; cerebral collaterals; arteriovenous malformations; ischemic limb angiogenesis; Osler-Webber Syndrome; plaque neovascularization; telangiectasia; hemophiliac joints; angiofibroma; fibromuscular dysplasia; wound granulation; Crohn's disease; and atherosclerosis.

For example, within one aspect of the present invention methods are provided for treating hypertrophic scars and keloids, comprising the step of administering a polynucleotide, polypeptide, antagonist and/or agonist of the invention to a hypertrophic scar or keloid.

Within one embodiment of the present invention polynucleotides, polypeptides, antagonists and/or agonists of the invention are directly injected into a hypertrophic scar or keloid, in order to prevent the progression of these lesions. This therapy is of particular value in the prophylactic treatment of conditions which are known to result in the development of hypertrophic scars and keloids (e.g., burns), and is preferably initiated after the proliferative phase has had time to progress (approximately 14 days after the initial injury), but before hypertrophic scar or keloid development. As noted above, the present invention also provides methods for treating neovascular diseases of the eye, including for example, corneal neovascularization, neovascular glaucoma, proliferative diabetic retinopathy, retrolental fibroplasia and macular degeneration.

Moreover, Ocular disorders associated with neovascularization which can be treated with the polynucleotides and polypeptides of the present invention (including agonists and/or antagonists) include, but are not limited to: neovascular glaucoma, diabetic retinopathy, retinoblastoma, retrolental fibroplasia, uveitis, retinopathy of prematurity macular degeneration, corneal graft neovascularization, as well as other eye inflammatory diseases, ocular tumors and diseases associated with choroidal or iris neovascularization. See, e.g., reviews by Waltman et al., Am. J. Ophthal. 85:704-710 (1978) and Gartner et al., Surv. Ophthal. 22:291-312 (1978).

Thus, within one aspect of the present invention methods are provided for treating neovascular diseases of the eye such as corneal neovascularization (including corneal graft neovascularization), comprising the step of administering to a patient a therapeutically effective amount of a compound (as described above) to the cornea, such that the formation of blood vessels is inhibited. Briefly, the cornea is a tissue that normally lacks blood vessels. In certain pathological conditions however, capillaries may extend into the cornea from the pericorneal vascular plexus of the limbus. When the cornea becomes vascularized, it also becomes clouded, resulting in a decline in the patient's visual acuity. Visual loss may become complete if the cornea completely opacitates. A wide variety of disorders can result in corneal neovascularization, including for example, corneal infections (e.g., trachoma, herpes simplex keratitis, leishmaniasis and onchocerciasis), immunological processes (e.g., graft rejection and Stevens-Johnson's syndrome), alkali burns, trauma, inflammation (of any cause), toxic and nutritional deficiency states, and as a complication of wearing contact lenses.

Within particularly preferred embodiments of the invention, may be prepared for topical administration in saline (combined with any of the preservatives and antimicrobial agents commonly used in ocular preparations), and administered in eyedrop form. The solution or suspension may be prepared in its pure form and administered several times daily. Alternatively, anti-angiogenic compositions, prepared as described above, may also be administered directly to the cornea. Within preferred embodiments, the anti-angiogenic composition is prepared with a muco-adhesive polymer that binds to cornea. Within further embodiments, the anti-angiogenic factors or anti-angiogenic compositions may be utilized as an adjunct to conventional steroid therapy. Topical therapy may also be useful prophylactically in corneal lesions which are known to have a high probability of inducing an angiogenic response (such as chemical burns). In these instances the treatment, likely in combination with steroids, may be instituted immediately to help prevent subsequent complications.

Within other embodiments, the compounds described above may be injected directly into the corneal stroma by an ophthalmologist under microscopic guidance. The preferred site of injection may vary with the morphology of the individual lesion, but the goal of the administration would be to place the composition at the advancing front of the vasculature (i.e., interspersed between the blood vessels and the normal cornea). In most cases this would involve perilimbic corneal injection to “protect” the cornea from the advancing blood vessels. This method may also be utilized shortly after a corneal insult in order to prophylactically prevent corneal neovascularization. In this situation the material could be injected in the perilimbic cornea interspersed between the corneal lesion and its undesired potential limbic blood supply. Such methods may also be utilized in a similar fashion to prevent capillary invasion of transplanted corneas. In a sustained-release form injections might only be required 2-3 times per year. A steroid could also be added to the injection solution to reduce inflammation resulting from the injection itself.

Within another aspect of the present invention, methods are provided for treating neovascular glaucoma, comprising the step of administering to a patient a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist to the eye, such that the formation of blood vessels is inhibited. In one embodiment, the compound may be administered topically to the eye in order to treat early forms of neovascular glaucoma. Within other embodiments, the compound may be implanted by injection into the region of the anterior chamber angle. Within other embodiments, the compound may also be placed in any location such that the compound is continuously released into the aqueous humor. Within another aspect of the present invention, methods are provided for treating proliferative diabetic retinopathy, comprising the step of administering to a patient a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist to the eyes, such that the formation of blood vessels is inhibited.

Within particularly preferred embodiments of the invention, proliferative diabetic retinopathy may be treated by injection into the aqueous humor or the vitreous, in order to increase the local concentration of the polynucleotide, polypeptide, antagonist and/or agonist in the retina. Preferably, this treatment should be initiated prior to the acquisition of severe disease requiring photocoagulation.

Within another aspect of the present invention, methods are provided for treating retrolental fibroplasia, comprising the step of administering to a patient a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist to the eye, such that the formation of blood vessels is inhibited. The compound may be administered topically, via intravitreous injection and/or via intraocular implants.

Additionally, disorders which can be treated with the polynucleotides, polypeptides, agonists and/or agonists include, but are not limited to, hemangioma, arthritis, psoriasis, angiofibroma, atherosclerotic plaques, delayed wound healing, granulations, hemophilic joints, hypertrophic scars, nonunion fractures, Osler-Weber syndrome, pyogenic granuloma, scleroderma, trachoma, and vascular adhesions.

Moreover, disorders and/or states, which can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated with the polynucleotides, polypeptides, agonists and/or agonists of the invention include, but are not limited to, solid tumors, blood born tumors such as leukemias, tumor metastasis, Kaposi's sarcoma, benign tumors, for example hemangiomas, acoustic neuromas, neurofibromas, trachomas, and pyogenic granulomas, rheumatoid arthritis, psoriasis, ocular angiogenic diseases, for example, diabetic retinopathy, retinopathy of prematurity, macular degeneration, corneal graft rejection, neovascular glaucoma, retrolental fibroplasia, rubeosis, retinoblastoma, and uvietis, delayed wound healing, endometriosis, vascluogenesis, granulations, hypertrophic scars (keloids), nonunion fractures, scleroderma, trachoma, vascular adhesions, myocardial angiogenesis, coronary collaterals, cerebral collaterals, arteriovenous malformations, ischemic limb angiogenesis, Osler-Webber Syndrome, plaque neovascularization, telangiectasia, hemophiliac joints, angiofibroma fibromuscular dysplasia, wound granulation, Crohn's disease, atherosclerosis, birth control agent by preventing vascularization required for embryo implantation controlling menstruation, diseases that have angiogenesis as a pathologic consequence such as cat scratch disease (Rochele minalia quintosa), ulcers (Helicobacter pylori), Bartonellosis and bacillary angiomatosis.

In one aspect of the birth control method, an amount of the compound sufficient to block embryo implantation is administered before or after intercourse and fertilization have occurred, thus providing an effective method of birth control, possibly a “morning after” method. Polynucleotides, polypeptides, agonists and/or agonists may also be used in controlling menstruation or administered as either a peritoneal lavage fluid or for peritoneal implantation in the treatment of endometriosis.

Polynucleotides, polypeptides, agonists and/or agonists of the present invention may be incorporated into surgical sutures in order to prevent stitch granulomas.

Polynucleotides, polypeptides, agonists and/or agonists may be utilized in a wide variety of surgical procedures. For example, within one aspect of the present invention a compositions (in the form of, for example, a spray or film) may be utilized to coat or spray an area prior to removal of a tumor, in order to isolate normal surrounding tissues from malignant tissue, and/or to prevent the spread of disease to surrounding tissues. Within other aspects of the present invention, compositions (e.g., in the form of a spray) may be delivered via endoscopic procedures in order to coat tumors, or inhibit angiogenesis in a desired locale. Within yet other aspects of the present invention, surgical meshes which have been coated with anti-angiogenic compositions of the present invention may be utilized in any procedure wherein a surgical mesh might be utilized. For example, within one embodiment of the invention a surgical mesh laden with an anti-angiogenic composition may be utilized during abdominal cancer resection surgery (e.g., subsequent to colon resection) in order to provide support to the structure, and to release an amount of the anti-angiogenic factor.

Within further aspects of the present invention, methods are provided for treating tumor excision sites, comprising administering a polynucleotide, polypeptide, agonist and/or agonist to the resection margins of a tumor subsequent to excision, such that the local recurrence of cancer and the formation of new blood vessels at the site is inhibited. Within one embodiment of the invention, the anti-angiogenic compound is administered directly to the tumor excision site (e.g., applied by swabbing, brushing or otherwise coating the resection margins of the tumor with the anti-angiogenic compound). Alternatively, the anti-angiogenic compounds may be incorporated into known surgical pastes prior to administration. Within particularly preferred embodiments of the invention, the anti-angiogenic compounds are applied after hepatic resections for malignancy, and after neurosurgical operations.

Within one aspect of the present invention, polynucleotides, polypeptides, agonists and/or agonists may be administered to the resection margin of a wide variety of tumors, including for example, breast, colon, brain and hepatic tumors. For example, within one embodiment of the invention, anti-angiogenic compounds may be administered to the site of a neurological tumor subsequent to excision, such that the formation of new blood vessels at the site are inhibited.

The polynucleotides, polypeptides, agonists and/or agonists of the present invention may also be administered along with other anti-angiogenic factors. Representative examples of other anti-angiogenic factors include: Anti-Invasive Factor, retinoic acid and derivatives thereof, paclitaxel, Suramin, Tissue Inhibitor of Metalloproteinase-1, Tissue Inhibitor of Metalloproteinase-2, Plasminogen Activator Inhibitor-1, Plasminogen Activator Inhibitor-2, and various forms of the lighter “d group” transition metals.

Lighter “d group” transition metals include, for example, vanadium, molybdenum, tungsten, titanium, niobium, and tantalum species. Such transition metal species may form transition metal complexes. Suitable complexes of the above-mentioned transition metal species include oxo transition metal complexes.

Representative examples of vanadium complexes include oxo vanadium complexes such as vanadate and vanadyl complexes. Suitable vanadate complexes include metavanadate and orthovanadate complexes such as, for example, ammonium metavanadate, sodium metavanadate, and sodium orthovanadate. Suitable vanadyl complexes include, for example, vanadyl acetylacetonate and vanadyl sulfate including vanadyl sulfate hydrates such as vanadyl sulfate mono- and trihydrates.

Representative examples of tungsten and molybdenum complexes also include oxo complexes. Suitable oxo tungsten complexes include tungstate and tungsten oxide complexes. Suitable tungstate complexes include ammonium tungstate, calcium tungstate, sodium tungstate dihydrate, and tungstic acid. Suitable tungsten oxides include tungsten (IV) oxide and tungsten (VI) oxide. Suitable oxo molybdenum complexes include molybdate, molybdenum oxide, and molybdenyl complexes. Suitable molybdate complexes include ammonium molybdate and its hydrates, sodium molybdate and its hydrates, and potassium molybdate and its hydrates. Suitable molybdenum oxides include molybdenum (VI) oxide, molybdenum (VI) oxide, and molybdic acid. Suitable molybdenyl complexes include, for example, molybdenyl acetylacetonate. Other suitable tungsten and molybdenum complexes include hydroxo derivatives derived from, for example, glycerol, tartaric acid, and sugars.

A wide variety of other anti-angiogenic factors may also be utilized within the context of the present invention. Representative examples include platelet factor 4; protamine sulphate; sulphated chitin derivatives (prepared from queen crab shells), (Murata et al., Cancer Res. 51:22-26, 1991); Sulphated Polysaccharide Peptidoglycan Complex (SP-PG) (the function of this compound may be enhanced by the presence of steroids such as estrogen, and tamoxifen citrate); Staurosporine; modulators of matrix metabolism, including for example, proline analogs, cishydroxyproline, d,L-3,4-dehydroproline, Thiaproline, alpha,alpha-dipyridyl, aminopropionitrile fumarate; 4-propyl-5-(4-pyridinyl)-2(3H)-oxazolone; Methotrexate; Mitoxantrone; Heparin; Interferons; 2 Macroglobulin-serum; ChIMP-3 (Pavloff et al., J. Bio. Chem. 267:17321-17326, 1992); Chymostatin (Tomkinson et al., Biochem J. 286:475-480, 1992); Cyclodextrin Tetradecasulfate; Eponemycin; Camptothecin; Fumagillin (Ingber et al., Nature 348:555-557, 1990); Gold Sodium Thiomalate (“GST”; Matsubara and Ziff, J. Clin. Invest. 79:1440-1446, 1987); anticollagenase-serum; alpha2-antiplasmin (Holmes et al., J. Biol. Chem. 262(4):1659-1664, 1987); Bisantrene (National Cancer Institute); Lobenzarit disodium (N-(2)-carboxyphenyl-4-chloroanthronilic acid disodium or “CCA”; Takeuchi et al., Agents Actions 36:312-316, 1992); Thalidomide; Angostatic steroid; AGM-1470; carboxynaminolmidazole; and metalloproteinase inhibitors such as BB94.

Diseases at the Cellular Level

Diseases associated with increased cell survival or the inhibition of apoptosis that could be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated using polynucleotides or polypeptides, as well as antagonists or agonists of the present invention, include cancers (such as follicular lymphomas, carcinomas with p53 mutations, and hormone-dependent tumors, including, but not limited to colon cancer, cardiac tumors, pancreatic cancer, melanoma, retinoblastoma, glioblastoma, lung cancer, intestinal cancer, testicular cancer, stomach cancer, neuroblastoma, myxoma, myoma, lymphoma, endothelioma, osteoblastoma, osteoclastoma, osteosarcoma, chondrosarcoma, adenoma, breast cancer, prostate cancer, Kaposi's sarcoma and ovarian cancer); autoimmune disorders (such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) and viral infections (such as herpes viruses, pox viruses and adenoviruses), inflammation, graft v. host disease, acute graft rejection, and chronic graft rejection.

In preferred embodiments, polynucleotides, polypeptides, and/or antagonists of the invention are used to inhibit growth, progression, and/or metasis of cancers, in particular those listed above.

Additional diseases or conditions associated with increased cell survival that could be treated or detected by polynucleotides or polypeptides, or agonists or antagonists of the present invention include, but are not limited to, progression, and/or metastases of malignancies and related disorders such as leukemia (including acute leukemias (e.g., acute lymphocytic leukemia, acute myelocytic leukemia (including myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia)) and chronic leukemias (e.g., chronic myelocytic (granulocytic) leukemia and chronic lymphocytic leukemia)), polycythemia vera, lymphomas (e.g., Hodgkin's disease and non-Hodgkin's disease), multiple myeloma, Waldenstrom's macroglobulinemia, heavy chain disease, and solid tumors including, but not limited to, sarcomas and carcinomas such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm's tumor, cervical cancer, testicular tumor, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, menangioma, melanoma, neuroblastoma, and retinoblastoma.

Diseases associated with increased apoptosis that could be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated using polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, include, but are not limited to, AIDS; neurodegenerative disorders (such as Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, Retinitis pigmentosa, Cerebellar degeneration and brain tumor or prior associated disease); autoimmune disorders (such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) myelodysplastic syndromes (such as aplastic anemia), graft v. host disease, ischemic injury (such as that caused by myocardial infarction, stroke and reperfusion injury), liver injury (e.g., hepatitis related liver injury, ischemia/reperfusion injury, cholestosis (bile duct injury) and liver cancer); toxin-induced liver disease (such as that caused by alcohol), septic shock, cachexia and anorexia.

Wound Healing and Epithelial Cell Proliferation

In accordance with yet a further aspect of the present invention, there is provided a process for utilizing polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, for therapeutic purposes, for example, to stimulate epithelial cell proliferation and basal keratinocytes for the purpose of wound healing, and to stimulate hair follicle production and healing of dermal wounds. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may be clinically useful in stimulating wound healing including surgical wounds, excisional wounds, deep wounds involving damage of the dermis and epidermis, eye tissue wounds, dental tissue wounds, oral cavity wounds, diabetic ulcers, dermal ulcers, cubitus ulcers, arterial ulcers, venous stasis ulcers, burns resulting from heat exposure or chemicals, and other abnormal wound healing conditions such as uremia, malnutrition, vitamin deficiencies and complications associated with systemic treatment with steroids, radiation therapy and antineoplastic drugs and antimetabolites. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to promote dermal reestablishment subsequent to dermal loss

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to increase the adherence of skin grafts to a wound bed and to stimulate re-epithelialization from the wound bed. The following are types of grafts that polynucleotides or polypeptides, agonists or antagonists of the present invention, could be used to increase adherence to a wound bed: autografts, artificial skin, allografts, autodermic graft, autoepdermic grafts, avacular grafts, Blair-Brown grafts, bone graft, brephoplastic grafts, cutis graft, delayed graft, dermic graft, epidermic graft, fascia graft, full thickness graft, heterologous graft, xenograft, homologous graft, hyperplastic graft, lamellar graft, mesh graft, mucosal graft, Ollier-Thiersch graft, omenpal graft, patch graft, pedicle graft, penetrating graft, split skin graft, thick split graft. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, can be used to promote skin strength and to improve the appearance of aged skin.

It is believed that polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, will also produce changes in hepatocyte proliferation, and epithelial cell proliferation in the lung, breast, pancreas, stomach, small intestine, and large intestine. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could promote proliferation of epithelial cells such as sebocytes, hair follicles, hepatocytes, type II pneumocytes, mucin-producing goblet cells, and other epithelial cells and their progenitors contained within the skin, lung, liver, and gastrointestinal tract. Polynucleotides or polypeptides, agonists or antagonists of the present invention, may promote proliferation of endothelial cells, keratinocytes, and basal keratinocytes.

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could also be used to reduce the side effects of gut toxicity that result from radiation, chemotherapy treatments or viral infections. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may have a cytoprotective effect on the small intestine mucosa. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may also stimulate healing of mucositis (mouth ulcers) that result from chemotherapy and viral infections.

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could further be used in full regeneration of skin in full and partial thickness skin defects, including burns, (i.e., repopulation of hair follicles, sweat glands, and sebaceous glands), treatment of other skin defects such as psoriasis. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to treat epidermolysis bullosa, a defect in adherence of the epidermis to the underlying dermis which results in frequent, open and painful blisters by accelerating reepithelialization of these lesions. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could also be used to treat gastric and doudenal ulcers and help heal by scar formation of the mucosal lining and regeneration of glandular mucosa and duodenal mucosal lining more rapidly. Inflammatory bowel diseases, such as Crohn's disease and ulcerative colitis, are diseases which result in destruction of the mucosal surface of the small or large intestine, respectively. Thus, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to promote the resurfacing of the mucosal surface to aid more rapid healing and to prevent progression of inflammatory bowel disease. Treatment with polynucleotides or polypeptides, agonists or antagonists of the present invention, is expected to have a significant effect on the production of mucus throughout the gastrointestinal tract and could be used to protect the intestinal mucosa from injurious substances that are ingested or following surgery. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to treat diseases associate with the under expression.

Moreover, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to prevent and heal damage to the lungs due to various pathological states. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, which could stimulate proliferation and differentiation and promote the repair of alveoli and brochiolar epithelium to prevent or treat acute or chronic lung damage. For example, emphysema, which results in the progressive loss of aveoli, and inhalation injuries, i.e., resulting from smoke inhalation and burns, that cause necrosis of the bronchiolar epithelium and alveoli could be effectively treated using polynucleotides or polypeptides, agonists or antagonists of the present invention. Also, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to stimulate the proliferation of and differentiation of type II pneumocytes, which may help treat or prevent disease such as hyaline membrane diseases, such as infant respiratory distress syndrome and bronchopulmonary displasia, in premature infants.

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could stimulate the proliferation and differentiation of hepatocytes and, thus, could be used to alleviate or treat liver diseases and pathologies such as fulminant liver failure caused by cirrhosis, liver damage caused by viral hepatitis and toxic substances (i.e., acetaminophen, carbon tetraholoride and other hepatotoxins known in the art).

In addition, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used treat or prevent the onset of diabetes mellitus. In patients with newly diagnosed Types I and II diabetes, where some islet cell function remains, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to maintain the islet function so as to alleviate, delay or prevent permanent manifestation of the disease. Also, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used as an auxiliary in islet cell transplantation to improve or promote islet cell function.

Neural Activity and Neurological Diseases

The polynucleotides, polypeptides and agonists or antagonists of the invention may be used for the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of diseases, disorders, damage or injury of the brain and/or nervous system. Nervous system disorders that can be treated with the compositions of the invention (e.g., polypeptides, polynucleotides, and/or agonists or antagonists), include, but are not limited to, nervous system injuries, and diseases or disorders which result in either a disconnection of axons, a diminution or degeneration of neurons, or demyelination. Nervous system lesions which may be treated in a patient (including human and non-human mammalian patients) according to the methods of the invention, include but are not limited to, the following lesions of either the central (including spinal cord, brain) or peripheral nervous systems: (1) ischemic lesions, in which a lack of oxygen in a portion of the nervous system results in neuronal injury or death, including cerebral infarction or ischemia, or spinal cord infarction or ischemia; (2) traumatic lesions, including lesions caused by physical injury or associated with surgery, for example, lesions which sever a portion of the nervous system, or compression injuries; (3) malignant lesions, in which a portion of the nervous system is destroyed or injured by malignant tissue which is either a nervous system associated malignancy or a malignancy derived from non-nervous system tissue; (4) infectious lesions, in which a portion of the nervous system is destroyed or injured as a result of infection, for example, by an abscess or associated with infection by human immunodeficiency virus, herpes zoster, or herpes simplex virus or with Lyme disease, tuberculosis, or syphilis; (5) degenerative lesions, in which a portion of the nervous system is destroyed or injured as a result of a degenerative process including but not limited to, degeneration associated with Parkinson's disease, Alzheimer's disease, Huntington's chorea, or amyotrophic lateral sclerosis (ALS); (6) lesions associated with nutritional diseases or disorders, in which a portion of the nervous system is destroyed or injured by a nutritional disorder or disorder of metabolism including, but not limited to, vitamin B12 deficiency, folic acid deficiency, Wernicke disease, tobacco-alcohol amblyopia, Marchiafava-Bignami disease (primary degeneration of the corpus callosum), and alcoholic cerebellar degeneration; (7) neurological lesions associated with systemic diseases including, but not limited to, diabetes (diabetic neuropathy, Bell's palsy), systemic lupus erythematosus, carcinoma, or sarcoidosis; (8) lesions caused by toxic substances including alcohol, lead, or particular neurotoxins; and (9) demyelinated lesions in which a portion of the nervous system is destroyed or injured by a demyelinating disease including, but not limited to, multiple sclerosis, human immunodeficiency virus-associated myelopathy, transverse myelopathy or various etiologies, progressive multifocal leukoencephalopathy, and central pontine myelinolysis.

In one embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to protect neural cells from the damaging effects of hypoxia. In a further preferred embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to protect neural cells from the damaging effects of cerebral hypoxia. According to this embodiment, the compositions of the invention are used to treat or prevent neural cell injury associated with cerebral hypoxia. In one non-exclusive aspect of this embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention, are used to treat or prevent neural cell injury associated with cerebral ischemia. In another non-exclusive aspect of this embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent neural cell injury associated with cerebral infarction.

In another preferred embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent neural cell injury associated with a stroke. In a specific embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent cerebral neural cell injury associated with a stroke.

In another preferred embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent neural cell injury associated with a heart attack. In a specific embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent cerebral neural cell injury associated with a heart attack.

The compositions of the invention which are useful for treating or preventing a nervous system disorder may be selected by testing for biological activity in promoting the survival or differentiation of neurons. For example, and not by way of limitation, compositions of the invention which elicit any of the following effects may be useful according to the invention: (1) increased survival time of neurons in culture either in the presence or absence of hypoxia or hypoxic conditions; (2) increased sprouting of neurons in culture or in vivo; (3) increased production of a neuron-associated molecule in culture or in vivo, e.g., choline acetyltransferase or acetylcholinesterase with respect to motor neurons; or (4) decreased symptoms of neuron dysfunction in vivo. Such effects may be measured by any method known in the art. In preferred, non-limiting embodiments, increased survival of neurons may routinely be measured using a method set forth herein or otherwise known in the art, such as, for example, in Zhang et al., Proc Natl Acad Sci USA 97:3637-42 (2000) or in Arakawa et al., J. Neurosci., 10:3507-15 (1990); increased sprouting of neurons may be detected by methods known in the art, such as, for example, the methods set forth in Pestronk et al., Exp. Neurol., 70:65-82 (1980), or Brown et al., Ann. Rev. Neurosci., 4:17-42 (1981); increased production of neuron-associated molecules may be measured by bioassay, enzymatic assay, antibody binding, Northern blot assay, etc., using techniques known in the art and depending on the molecule to be measured; and motor neuron dysfunction may be measured by assessing the physical manifestation of motor neuron disorder, e.g., weakness, motor neuron conduction velocity, or functional disability.

In specific embodiments, motor neuron disorders that may be treated according to the invention include, but are not limited to, disorders such as infarction, infection, exposure to toxin, trauma, surgical damage, degenerative disease or malignancy that may affect motor neurons as well as other components of the nervous system, as well as disorders that selectively affect neurons such as amyotrophic lateral sclerosis, and including, but not limited to, progressive spinal muscular atrophy, progressive bulbar palsy, primary lateral sclerosis, infantile and juvenile muscular atrophy, progressive bulbar paralysis of childhood (Fazio-Londe syndrome), poliomyelitis and the post polio syndrome, and Hereditary Motorsensory Neuropathy (Charcot-Marie-Tooth Disease).

Further, polypeptides or polynucleotides of the invention may play a role in neuronal survival; synapse formation; conductance; neural differentiation, etc. Thus, compositions of the invention (including polynucleotides, polypeptides, and agonists or antagonists) may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate diseases or disorders associated with these roles, including, but not limited to, learning and/or cognition disorders. The compositions of the invention may also be useful in the treatment or prevention of neurodegenerative disease states and/or behavioural disorders. Such neurodegenerative disease states and/or behavioral disorders include, but are not limited to, Alzheimer's Disease, Parkinson's Disease, Huntington's Disease, Tourette Syndrome, schizophrenia, mania, dementia, paranoia, obsessive compulsive disorder, panic disorder, learning disabilities, ALS, psychoses, autism, and altered behaviors, including disorders in feeding, sleep patterns, balance, and perception. In addition, compositions of the invention may also play a role in the treatment, prevention and/or detection of developmental disorders associated with the developing embryo, or sexually-linked disorders.

Additionally, polypeptides, polynucleotides and/or agonists or antagonists of the invention, may be useful in protecting neural cells from diseases, damage, disorders, or injury, associated with cerebrovascular disorders including, but not limited to, carotid artery diseases (e.g., carotid artery thrombosis, carotid stenosis, or Moyamoya Disease), cerebral amyloid angiopathy, cerebral aneurysm, cerebral anoxia, cerebral arteriosclerosis, cerebral arteriovenous malformations, cerebral artery diseases, cerebral embolism and thrombosis (e.g., carotid artery thrombosis, sinus thrombosis, or Wallenberg's Syndrome), cerebral hemorrhage (e.g., epidural or subdural hematoma, or subarachnoid hemorrhage), cerebral infarction, cerebral ischemia (e.g., transient cerebral ischemia, Subclavian Steal Syndrome, or vertebrobasilar insufficiency), vascular dementia (e.g., multi-infarct), leukomalacia, periventricular, and vascular headache (e.g., cluster headache or migraines).

In accordance with yet a further aspect of the present invention, there is provided a process for utilizing polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, for therapeutic purposes, for example, to stimulate neurological cell proliferation and/or differentiation. Therefore, polynucleotides, polypeptides, agonists and/or antagonists of the invention may be used to treat and/or detect neurologic diseases. Moreover, polynucleotides or polypeptides, or agonists or antagonists of the invention, can be used as a marker or detector of a particular nervous system disease or disorder.

Examples of neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include brain diseases, such as metabolic brain diseases which includes phenylketonuria such as maternal phenylketonuria, pyruvate carboxylase deficiency, pyruvate dehydrogenase complex deficiency, Wernicke's Encephalopathy, brain edema, brain neoplasms such as cerebellar neoplasms which include infratentorial neoplasms, cerebral ventricle neoplasms such as choroid plexus neoplasms, hypothalamic neoplasms, supratentorial neoplasms, canavan disease, cerebellar diseases such as cerebellar ataxia which include spinocerebellar degeneration such as ataxia telangiectasia, cerebellar dyssynergia, Friederich's Ataxia, Machado-Joseph Disease, olivopontocerebellar atrophy, cerebellar neoplasms such as infratentorial neoplasms, diffuse cerebral sclerosis such as encephalitis periaxialis, globoid cell leukodystrophy, metachromatic leukodystrophy and subacute sclerosing panencephalitis.

Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include cerebrovascular disorders (such as carotid artery diseases which include carotid artery thrombosis, carotid stenosis and Moyamoya Disease), cerebral amyloid angiopathy, cerebral aneurysm, cerebral anoxia, cerebral arteriosclerosis, cerebral arteriovenous malformations, cerebral artery diseases, cerebral embolism and thrombosis such as carotid artery thrombosis, sinus thrombosis and Wallenberg's Syndrome, cerebral hemorrhage such as epidural hematoma, subdural hematoma and subarachnoid hemorrhage, cerebral infarction, cerebral ischemia such as transient cerebral ischemia, Subclavian Steal Syndrome and vertebrobasilar insufficiency, vascular dementia such as multi-infarct dementia, periventricular leukomalacia, vascular headache such as cluster headache and migraine.

Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include dementia such as AIDS Dementia Complex, presenile dementia such as Alzheimer's Disease and Creutzfeldt-Jakob Syndrome, senile dementia such as Alzheimer's Disease and progressive supranuclear palsy, vascular dementia such as multi-infarct dementia, encephalitis which include encephalitis periaxialis, viral encephalitis such as epidemic encephalitis, Japanese Encephalitis, St. Louis Encephalitis, tick-borne encephalitis and West Nile Fever, acute disseminated encephalomyelitis, meningoencephalitis such as uveomeningoencephalitic syndrome, Postencephalitic Parkinson Disease and subacute sclerosing panencephalitis, encephalomalacia such as periventricular leukomalacia, epilepsy such as generalized epilepsy which includes infantile spasms, absence epilepsy, myoclonic epilepsy which includes MERRF Syndrome, tonic-clonic epilepsy, partial epilepsy such as complex partial epilepsy, frontal lobe epilepsy and temporal lobe epilepsy, post-traumatic epilepsy, status epilepticus such as Epilepsia Partialis Continua, and Hallervorden-Spatz Syndrome.

Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include hydrocephalus such as Dandy-Walker Syndrome and normal pressure hydrocephalus, hypothalamic diseases such as hypothalamic neoplasms, cerebral malaria, narcolepsy which includes cataplexy, bulbar poliomyelitis, cerebri pseudotumor, Rett Syndrome, Reye's Syndrome, thalamic diseases, cerebral toxoplasmosis, intracranial tuberculoma and Zellweger Syndrome, central nervous system infections such as AIDS Dementia Complex, Brain Abscess, subdural empyema, encephalomyelitis such as Equine Encephalomyelitis, Venezuelan Equine Encephalomyelitis, Necrotizing Hemorrhagic Encephalomyelitis, Visna, and cerebral malaria.

Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include meningitis such as arachnoiditis, aseptic meningtitis such as viral meningtitis which includes lymphocytic choriomeningitis, Bacterial meningtitis which includes Haemophilus Meningtitis, Listeria Meningtitis, Meningococcal Meningtitis such as Waterhouse-Friderichsen Syndrome, Pneumococcal Meningtitis and meningeal tuberculosis, fungal meningitis such as Cryptococcal Meningtitis, subdural effusion, meningoencephalitis such as uvemeningoencephalitic syndrome, myelitis such as transverse myelitis, neurosyphilis such as tabes dorsalis, poliomyelitis which includes bulbar poliomyelitis and postpoliomyelitis syndrome, prion diseases (such as Creutzfeldt-Jakob Syndrome, Bovine Spongiform Encephalopathy, Gerstmann-Straussler Syndrome, Kuru, Scrapie), and cerebral toxoplasmosis.

Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include central nervous system neoplasms such as brain neoplasms that include cerebellar neoplasms such as infratentorial neoplasms, cerebral ventricle neoplasms such as choroid plexus neoplasms, hypothalamic neoplasms and supratentorial neoplasms, meningeal neoplasms, spinal cord neoplasms which include epidural neoplasms, demyelinating diseases such as Canavan Diseases, diffuse cerebral sceloris which includes adrenoleukodystrophy, encephalitis periaxialis, globoid cell leukodystrophy, diffuse cerebral sclerosis such as metachromatic leukodystrophy, allergic encephalomyelitis, necrotizing hemorrhagic encephalomyelitis, progressive multifocal leukoencephalopathy, multiple sclerosis, central pontine myelinolysis, transverse myelitis, neuromyelitis optica, Scrapie, Swayback, Chronic Fatigue Syndrome, Visna, High Pressure Nervous Syndrome, Meningism, spinal cord diseases such as amyotonia congenita, amyotrophic lateral sclerosis, spinal muscular atrophy such as Werdnig-Hoffmann Disease, spinal cord compression, spinal cord neoplasms such as epidural neoplasms, syringomyelia, Tabes Dorsalis, Stiff-Man Syndrome, mental retardation such as Angelman Syndrome, Cri-du-Chat Syndrome, De Lange's Syndrome, Down Syndrome, Gangliosidoses such as gangliosidoses G(M1), Sandhoff Disease, Tay-Sachs Disease, Hartnup Disease, homocystinuria, Laurence-Moon-Biedl Syndrome, Lesch-Nyhan Syndrome, Maple Syrup Urine Disease, mucolipidosis such as fucosidosis, neuronal ceroid-lipofuscinosis, oculocerebrorenal syndrome, phenylketonuria such as maternal phenylketonuria, Prader-Willi Syndrome, Rett Syndrome, Rubinstein-Taybi Syndrome, Tuberous Sclerosis, WAGR Syndrome, nervous system abnormalities such as holoprosencephaly, neural tube defects such as anencephaly which includes hydrangencephaly, Arnold-Chain Deformity, encephalocele, meningocele, meningomyelocele, spinal dysraphism such as spina bifida cystica and spina bifida occulta.

Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include hereditary motor and sensory neuropathies which include Charcot-Marie Disease, Hereditary optic atrophy, Refsum's Disease, hereditary spastic paraplegia, Werdnig-Hoffmann Disease, Hereditary Sensory and Autonomic Neuropathies such as Congenital Analgesia and Familial Dysautonomia, Neurologic manifestations (such as agnosia that include Gerstmann's Syndrome, Amnesia such as retrograde amnesia, apraxia, neurogenic bladder, cataplexy, communicative disorders such as hearing disorders that includes deafness, partial hearing loss, loudness recruitment and tinnitus, language disorders such as aphasia which include agraphia, anomia, broca aphasia, and Wernicke Aphasia, Dyslexia such as Acquired Dyslexia, language development disorders, speech disorders such as aphasia which includes anomia, broca aphasia and Wernicke Aphasia, articulation disorders, communicative disorders such as speech disorders which include dysarthria, echolalia, mutism and stuttering, voice disorders such as aphonia and hoarseness, decerebrate state, delirium, fasciculation, hallucinations, meningism, movement disorders such as angelman syndrome, ataxia, athetosis, chorea, dystonia, hypokinesia, muscle hypotonia, myoclonus, tic, torticollis and tremor, muscle hypertonia such as muscle rigidity such as stiff-man syndrome, muscle spasticity, paralysis such as facial paralysis which includes Herpes Zoster Oticus, Gastroparesis, Hemiplegia, ophthalmoplegia such as diplopia, Duane's Syndrome, Horner's Syndrome, Chronic progressive external ophthalmoplegia such as Kearns Syndrome, Bulbar Paralysis, Tropical Spastic Paraparesis, Paraplegia such as Brown-Sequard Syndrome, quadriplegia, respiratory paralysis and vocal cord paralysis, paresis, phantom limb, taste disorders such as ageusia and dysgeusia, vision disorders such as amblyopia, blindness, color vision defects, diplopia, hemianopsia, scotoma and subnormal vision, sleep disorders such as hypersomnia which includes Kleine-Levin Syndrome, insomnia, and somnambulism, spasm such as trismus, unconsciousness such as coma, persistent vegetative state and syncope and vertigo, neuromuscular diseases such as amyotonia congenita, amyotrophic lateral sclerosis, Lambert-Eaton Myasthenic Syndrome, motor neuron disease, muscular atrophy such as spinal muscular atrophy, Charcot-Marie Disease and Werdnig-Hoffmann Disease, Postpoliomyelitis Syndrome, Muscular Dystrophy, Myasthenia Gravis, Myotonia Atrophica, Myotonia Confenita, Nemaline Myopathy, Familial Periodic Paralysis, Multiplex Paramyloclonus, Tropical Spastic Paraparesis and Stiff-Man Syndrome, peripheral nervous system diseases such as acrodynia, amyloid neuropathies, autonomic nervous system diseases such as Adie's Syndrome, Barre-Lieou Syndrome, Familial Dysautonomia, Horner's Syndrome, Reflex Sympathetic Dystrophy and Shy-Drager Syndrome, Cranial Nerve Diseases such as Acoustic Nerve Diseases such as Acoustic Neuroma which includes Neurofibromatosis 2, Facial Nerve Diseases such as Facial Neuralgia, Melkersson-Rosenthal Syndrome, ocular motility disorders which includes amblyopia, nystagmus, oculomotor nerve paralysis, ophthalmoplegia such as Duane's Syndrome, Horner's Syndrome, Chronic Progressive External Ophthalmoplegia which includes Kearns Syndrome, Strabismus such as Esotropia and Exotropia, Oculomotor Nerve Paralysis, Optic Nerve Diseases such as Optic Atrophy which includes Hereditary Optic Atrophy, Optic Disk Drusen, Optic Neuritis such as Neuromyelitis Optica, Papilledema, Trigeminal Neuralgia, Vocal Cord Paralysis, Demyelinating Diseases such as Neuromyelitis Optica and Swayback, and Diabetic neuropathies such as diabetic foot.

Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include nerve compression syndromes such as carpal tunnel syndrome, tarsal tunnel syndrome, thoracic outlet syndrome such as cervical rib syndrome, ulnar nerve compression syndrome, neuralgia such as causalgia, cervico-brachial neuralgia, facial neuralgia and trigeminal neuralgia, neuritis such as experimental allergic neuritis, optic neuritis, polyneuritis, polyradiculoneuritis and radiculities such as polyradiculitis, hereditary motor and sensory neuropathies such as Charcot-Marie Disease, Hereditary Optic Atrophy, Refsum's Disease, Hereditary Spastic Paraplegia and Werdnig-Hoffmann Disease, Hereditary Sensory and Autonomic Neuropathies which include Congenital Analgesia and Familial Dysautonomia, POEMS Syndrome, Sciatica, Gustatory Sweating and Tetany).

Endocrine Disorders

Polynucleotides or polypeptides, or agonists or antagonists of the present invention, may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate disorders and/or diseases related to hormone imbalance, and/or disorders or diseases of the endocrine system.

Hormones secreted by the glands of the endocrine system control physical growth, sexual function, metabolism, and other functions. Disorders may be classified in two ways: disturbances in the production of hormones, and the inability of tissues to respond to hormones. The etiology of these hormone imbalance or endocrine system diseases, disorders or conditions may be genetic, somatic, such as cancer and some autoimmune diseases, acquired (e.g., by chemotherapy, injury or toxins), or infectious. Moreover, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention can be used as a marker or detector of a particular disease or disorder related to the endocrine system and/or hormone imbalance.

Endocrine system and/or hormone imbalance and/or diseases encompass disorders of uterine motility including, but not limited to: complications with pregnancy and labor (e.g., pre-term labor, post-term pregnancy, spontaneous abortion, and slow or stopped labor); and disorders and/or diseases of the menstrual cycle (e.g., dysmenorrhea and endometriosis).

Endocrine system and/or hormone imbalance disorders and/or diseases include disorders and/or diseases of the pancreas, such as, for example, diabetes mellitus, diabetes insipidus, congenital pancreatic agenesis, pheochromocytoma—islet cell tumor syndrome; disorders and/or diseases of the adrenal glands such as, for example, Addison's Disease, corticosteroid deficiency, virilizing disease, hirsutism, Cushing's Syndrome, hyperaldosteronism, pheochromocytoma; disorders and/or diseases of the pituitary gland, such as, for example, hyperpituitarism, hypopituitarism, pituitary dwarfism, pituitary adenoma, panhypopituitarism, acromegaly, gigantism; disorders and/or diseases of the thyroid, including but not limited to, hyperthyroidism, hypothyroidism, Plummer's disease, Graves' disease (toxic diffuse goiter), toxic nodular goiter, thyroiditis (Hashimoto's thyroiditis, subacute granulomatous thyroiditis, and silent lymphocytic thyroiditis), Pendred's syndrome, myxedema, cretinism, thyrotoxicosis, thyroid hormone coupling defect, thymic aplasia, Hurthle cell tumours of the thyroid, thyroid cancer, thyroid carcinoma, Medullary thyroid carcinoma; disorders and/or diseases of the parathyroid, such as, for example, hyperparathyroidism, hypoparathyroidism; disorders and/or diseases of the hypothalamus.

In addition, endocrine system and/or hormone imbalance disorders and/or diseases may also include disorders and/or diseases of the testes or ovaries, including cancer. Other disorders and/or diseases of the testes or ovaries further include, for example, ovarian cancer, polycystic ovary syndrome, Klinefelter's syndrome, vanishing testes syndrome (bilateral anorchia), congenital absence of Leydig's cells, cryptorchidism, Noonan's syndrome, myotonic dystrophy, capillary haemangioma of the testis (benign), neoplasias of the testis and neo-testis.

Moreover, endocrine system and/or hormone imbalance disorders and/or diseases may also include disorders and/or diseases such as, for example, polyglandular deficiency syndromes, pheochromocytoma, neuroblastoma, multiple Endocrine neoplasia, and disorders and/or cancers of endocrine tissues.

In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate endocrine diseases and/or disorders associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1B.2, column 5 (Tissue Distribution Library Code).

Reproductive System Disorders

The polynucleotides or polypeptides, or agonists or antagonists of the invention may be used for the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of diseases and/or disorders of the reproductive system. Reproductive system disorders that can be treated by the compositions of the invention, include, but are not limited to, reproductive system injuries, infections, neoplastic disorders, congenital defects, and diseases or disorders which result in infertility, complications with pregnancy, labor, or parturition, and postpartum difficulties.

Reproductive system disorders and/or diseases include diseases and/or disorders of the testes, including testicular atrophy, testicular feminization, cryptorchism (unilateral and bilateral), anorchia, ectopic testis, epididymitis and orchitis (typically resulting from infections such as, for example, gonorrhea, mumps, tuberculosis, and syphilis), testicular torsion, vasitis nodosa, germ cell tumors (e.g., seminomas, embryonal cell carcinomas, teratocarcinomas, choriocarcinomas, yolk sac tumors, and teratomas), stromal tumors (e.g., Leydig cell tumors), hydrocele, hematocele, varicocele, spermatocele, inguinal hernia, and disorders of sperm production (e.g., immotile cilia syndrome, aspermia, asthenozoospermia, azoospermia, oligospermia, and teratozoospermia).

Reproductive system disorders also include disorders of the prostate gland, such as acute non-bacterial prostatitis, chronic non-bacterial prostatitis, acute bacterial prostatitis, chronic bacterial prostatitis, prostatodystonia, prostatosis, granulomatous prostatitis, malacoplakia, benign prostatic hypertrophy or hyperplasia, and prostate neoplastic disorders, including adenocarcinomas, transitional cell carcinomas, ductal carcinomas, and squamous cell carcinomas.

Additionally, the compositions of the invention may be useful in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of disorders or diseases of the penis and urethra, including inflammatory disorders, such as balanoposthitis, balanitis xerotica obliterans, phimosis, paraphimosis, syphilis, herpes simplex virus, gonorrhea, non-gonococcal urethritis, chlamydia, mycoplasma, trichomonas, HIV, AIDS, Reiter's syndrome, condyloma acuminatum, condyloma latum, and pearly penile papules; urethral abnormalities, such as hypospadias, epispadias, and phimosis; premalignant lesions, including Erythroplasia of Queyrat, Bowen's disease, Bowenoid paplosis, giant condyloma of Buscke-Lowenstein, and varrucous carcinoma; penile cancers, including squamous cell carcinomas, carcinoma in situ, verrucous carcinoma, and disseminated penile carcinoma; urethral neoplastic disorders, including penile urethral carcinoma, bulbomembranous urethral carcinoma, and prostatic urethral carcinoma; and erectile disorders, such as priapism, Peyronie's disease, erectile dysfunction, and impotence.

Moreover, diseases and/or disorders of the vas deferens include vasculititis and CBAVD (congenital bilateral absence of the vas deferens); additionally, the polynucleotides, polypeptides, and agonists or antagonists of the present invention may be used in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of diseases and disorders of the seminal vesicles, including hydatid disease, congenital chloride diarrhea, and polycystic kidney disease.

Other disorders and/or diseases of the male reproductive system include, for example, Klinefelter's syndrome, Young's syndrome, premature ejaculation, diabetes mellitus, cystic fibrosis, Kartagener's syndrome, high fever, multiple sclerosis, and gynecomastia.

Further, the polynucleotides, polypeptides, and agonists or antagonists of the present invention may be used in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of diseases and/or disorders of the vagina and vulva, including bacterial vaginosis, candida vaginitis, herpes simplex virus, chancroid, granuloma inguinale, lymphogranuloma venereum, scabies, human papillomavirus, vaginal trauma, vulvar trauma, adenosis, chlamydia vaginitis, gonorrhea, trichomonas vaginitis, condyloma acuminatum, syphilis, molluscum contagiosum, atrophic vaginitis, Paget's disease, lichen sclerosus, lichen planus, vulvodynia, toxic shock syndrome, vaginismus, vulvovaginitis, vulvar vestibulitis, and neoplastic disorders, such as squamous cell hyperplasia, clear cell carcinoma, basal cell carcinoma, melanomas, cancer of Bartholin's gland, and vulvar intraepithelial neoplasia.

Disorders and/or diseases of the uterus include dysmenorrhea, retroverted uterus, endometriosis, fibroids, adenomyosis, anovulatory bleeding, amenorrhea, Cushing's syndrome, hydatidiform moles, Asherman's syndrome, premature menopause, precocious puberty, uterine polyps, dysfunctional uterine bleeding (e.g., due to aberrant hormonal signals), and neoplastic disorders, such as adenocarcinomas, keiomyosarcomas, and sarcomas. Additionally, the polypeptides, polynucleotides, or agonists or antagonists of the invention may be useful as a marker or detector of, as well as in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of congenital uterine abnormalities, such as bicornuate uterus, septate uterus, simple unicornuate uterus, unicornuate uterus with a noncavitary rudimentary horn, unicornuate uterus with a non-communicating cavitary rudimentary horn, unicornuate uterus with a communicating cavitary horn, arcuate uterus, uterine didelfus, and T-shaped uterus.

Ovarian diseases and/or disorders include anovulation, polycystic ovary syndrome (Stein-Leventhal syndrome), ovarian cysts, ovarian hypofunction, ovarian insensitivity to gonadotropins, ovarian overproduction of androgens, right ovarian vein syndrome, amenorrhea, hirutism, and ovarian cancer (including, but not limited to, primary and secondary cancerous growth, Sertoli-Leydig tumors, endometriod carcinoma of the ovary, ovarian papillary serous adenocarcinoma, ovarian mucinous adenocarcinoma, and Ovarian Krukenberg tumors).

Cervical diseases and/or disorders include cervicitis, chronic cervicitis, mucopurulent cervicitis, cervical dysplasia, cervical polyps, Nabothian cysts, cervical erosion, cervical incompetence, and cervical neoplasms (including, for example, cervical carcinoma, squamous metaplasia, squamous cell carcinoma, adenosquamous cell neoplasia, and columnar cell neoplasia).

Additionally, diseases and/or disorders of the reproductive system include disorders and/or diseases of pregnancy, including miscarriage and stillbirth, such as early abortion, late abortion, spontaneous abortion, induced abortion, therapeutic abortion, threatened abortion, missed abortion, incomplete abortion, complete abortion, habitual abortion, missed abortion, and septic abortion; ectopic pregnancy, anemia, Rh incompatibility, vaginal bleeding during pregnancy, gestational diabetes, intrauterine growth retardation, polyhydramnios, HELLP syndrome, abruptio placentae, placenta previa, hyperemesis, preeclampsia, eclampsia, herpes gestationis, and urticaria of pregnancy. Additionally, the polynucleotides, polypeptides, and agonists or antagonists of the present invention may be used in the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of diseases that can complicate pregnancy, including heart disease, heart failure, rheumatic heart disease, congenital heart disease, mitral valve prolapse, high blood pressure, anemia, kidney disease, infectious disease (e.g., rubella, cytomegalovirus, toxoplasmosis, infectious hepatitis, chlamydia, HIV, AIDS, and genital herpes), diabetes mellitus, Graves' disease, thyroiditis, hypothyroidism, Hashimoto's thyroiditis, chronic active hepatitis, cirrhosis of the liver, primary biliary cirrhosis, asthma, systemic lupus eryematosis, rheumatoid arthritis, myasthenia gravis, idiopathic thrombocytopenic purpura, appendicitis, ovarian cysts, gallbladder disorders, and obstruction of the intestine.

Complications associated with labor and parturition include premature rupture of the membranes, pre-term labor, post-term pregnancy, postmaturity, labor that progresses too slowly, fetal distress (e.g., abnormal heart rate (fetal or maternal), breathing problems, and abnormal fetal position), shoulder dystocia, prolapsed umbilical cord, amniotic fluid embolism, and aberrant uterine bleeding.

Further, diseases and/or disorders of the postdelivery period, including endometritis, myometritis, parametritis, peritonitis, pelvic thrombophlebitis, pulmonary embolism, endotoxemia, pyelonephritis, saphenous thrombophlebitis, mastitis, cystitis, postpartum hemorrhage, and inverted uterus.

Other disorders and/or diseases of the female reproductive system that may be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by the polynucleotides, polypeptides, and agonists or antagonists of the present invention include, for example, Turner's syndrome, pseudohermaphroditism, premenstrual syndrome, pelvic inflammatory disease, pelvic congestion (vascular engorgement), frigidity, anorgasmia, dyspareunia, ruptured fallopian tube, and Mittelschmerz.

Infectious Disease

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention can be used to treat or detect infectious agents. For example, by increasing the immune response, particularly increasing the proliferation and differentiation of B and/or T cells, infectious diseases may be treated. The immune response may be increased by either enhancing an existing immune response, or by initiating a new immune response. Alternatively, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may also directly inhibit the infectious agent, without necessarily eliciting an immune response.

Viruses are one example of an infectious agent that can cause disease or symptoms that can be treated or detected by a polynucleotide or polypeptide and/or agonist or antagonist of the present invention. Examples of viruses, include, but are not limited to Examples of viruses, include, but are not limited to the following DNA and RNA viruses and viral families: Arbovirus, Adenoviridae, Arenaviridae, Arterivirus, Birnaviridae, Bunyaviridae, Caliciviridae, Circoviridae, Coronaviridae, Dengue, EBV, HIV, Flaviviridae, Hepadnaviridae (Hepatitis), Herpesviridae (such as, Cytomegalovirus, Herpes Simplex, Herpes Zoster), Mononegavirus (e.g., Paramyxoviridae, Morbillivirus, Rhabdoviridae), Orthomyxoviridae (e.g., Influenza A, Influenza B, and parainfluenza), Papiloma virus, Papovaviridae, Parvoviridae, Picornaviridae, Poxyiridae (such as Smallpox or Vaccinia), Reoviridae (e.g., Rotavirus), Retroviridae (HTLV-I, HTLV-II, Lentivirus), and Togaviridae (e.g., Rubivirus). Viruses falling within these families can cause a variety of diseases or symptoms, including, but not limited to: arthritis, bronchiollitis, respiratory syncytial virus, encephalitis, eye infections (e.g., conjunctivitis, keratitis), chronic fatigue syndrome, hepatitis (A, B, C, E, Chronic Active, Delta), Japanese B encephalitis, Junin, Chikungunya, Rift Valley fever, yellow fever, meningitis, opportunistic infections (e.g., AIDS), pneumonia, Burkitt's Lymphoma, chickenpox, hemorrhagic fever, Measles, Mumps, Parainfluenza, Rabies, the common cold, Polio, leukemia, Rubella, sexually transmitted diseases, skin diseases (e.g., Kaposi's, warts), and viremia. polynucleotides or polypeptides, or agonists or antagonists of the invention, can be used to treat or detect any of these symptoms or diseases. In specific embodiments, polynucleotides, polypeptides, or agonists or antagonists of the invention are used to treat: meningitis, Dengue, EBV, and/or hepatitis (e.g., hepatitis B). In an additional specific embodiment polynucleotides, polypeptides, or agonists or antagonists of the invention are used to treat patients nonresponsive to one or more other commercially available hepatitis vaccines. In a further specific embodiment polynucleotides, polypeptides, or agonists or antagonists of the invention are used to treat AIDS.

Similarly, bacterial and fungal agents that can cause disease or symptoms and that can be treated or detected by a polynucleotide or polypeptide and/or agonist or antagonist of the present invention include, but not limited to, the following Gram-Negative and Gram-positive bacteria, bacterial families, and fungi: Actinomyces (e.g., Norcardia), Acinetobacter, Cryptococcus neoformans, Aspergillus, Bacillaceae (e.g., Bacillus anthrasis), Bacteroides (e.g., Bacteroides fragilis), Blastomycosis, Bordetella, Borrelia (e.g., Borrelia burgdorferi), Brucella, Candidia, Campylobacter, Chlamydia, Clostridium (e.g., Clostridium botulinum, Clostridium dificile, Clostridium perfringens, Clostridium tetani), Coccidioides, Corynebacterium (e.g., Corynebacterium diptheriae), Cryptococcus, Dermatocycoses, E. coli (e.g., Enterotoxigenic E. coli and Enterohemorrhagic E. coli), Enterobacter (e.g. Enterobacter aerogenes), Enterobacteriaceae (Klebsiella, Salmonella (e.g., Salmonella typhi, Salmonella enteritidis, Salmonella typhi), Serratia, Yersinia, Shigella), Erysipelothrix, Haemophilus (e.g., Haemophilus influenza type B), Helicobacter, Legionella (e.g., Legionella pneumophila), Leptospira, Listeria (e.g., Listeria monocytogenes), Mycoplasma, Mycobacterium (e.g., Mycobacterium leprae and Mycobacterium tuberculosis), Vibrio (e.g., Vibrio cholerae), Neisseriaceae (e.g., Neisseria gonorrhea, Neisseria meningitidis), Pasteurellacea, Proteus, Pseudomonas (e.g., Pseudomonas aeruginosa), Rickettsiaceae, Spirochetes (e.g., Treponema spp., Leptospira spp., Borrelia spp.), Shigella spp., Staphylococcus (e.g., Staphylococcus aureus), Meningiococcus, Pneumococcus and Streptococcus (e.g., Streptococcus pneumoniae and Groups A, B, and C Streptococci), and Ureaplasmas. These bacterial, parasitic, and fungal families can cause diseases or symptoms, including, but not limited to: antibiotic-resistant infections, bacteremia, endocarditis, septicemia, eye infections (e.g., conjunctivitis), uveitis, tuberculosis, gingivitis, bacterial diarrhea, opportunistic infections (e.g., AIDS related infections), paronychia, prosthesis-related infections, dental caries, Reiter's Disease, respiratory tract infections, such as Whooping Cough or Empyema, sepsis, Lyme Disease, Cat-Scratch Disease, dysentery, paratyphoid fever, food poisoning, Legionella disease, chronic and acute inflammation, erythema, yeast infections, typhoid, pneumonia, gonorrhea, meningitis (e.g., mengitis types A and B), chlamydia, syphillis, diphtheria, leprosy, brucellosis, peptic ulcers, anthrax, spontaneous abortions, birth defects, pneumonia, lung infections, ear infections, deafness, blindness, lethargy, malaise, vomiting, chronic diarrhea, Crohn's disease, colitis, vaginosis, sterility, pelvic inflammatory diseases, candidiasis, paratuberculosis, tuberculosis, lupus, botulism, gangrene, tetanus, impetigo, Rheumatic Fever, Scarlet Fever, sexually transmitted diseases, skin diseases (e.g., cellulitis, dermatocycoses), toxemia, urinary tract infections, wound infections, noscomial infections. Polynucleotides or polypeptides, agonists or antagonists of the invention, can be used to treat or detect any of these symptoms or diseases. In specific embodiments, polynucleotides, polypeptides, agonists or antagonists of the invention are used to treat: tetanus, diptheria, botulism, and/or meningitis type B.

Moreover, parasitic agents causing disease or symptoms that can be detected, prevented, diagnosed, prognosticated, treated, and/or ameliorated by a polynucleotide or polypeptide and/or agonist or antagonist of the present invention include, but not limited to, the following families or class: Amebiasis, Babesiosis, Coccidiosis, Cryptosporidiosis, Dientamoebiasis, Dourine, Ectoparasitic, Giardias, Helminthiasis, Leishmaniasis, Schistisoma, Theileriasis, Toxoplasmosis, Trypanosomiasis, and Trichomonas and Sporozoans (e.g., Plasmodium virax, Plasmodium falciparium, Plasmodium malariae and Plasmodium ovale). These parasites can cause a variety of diseases or symptoms, including, but not limited to: Scabies, Trombiculiasis, eye infections, intestinal disease (e.g., dysentery, giardiasis), liver disease, lung disease, opportunistic infections (e.g., AIDS related), malaria, pregnancy complications, and toxoplasmosis. polynucleotides or polypeptides, or agonists or antagonists of the invention, can be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate any of these symptoms or diseases. In specific embodiments, polynucleotides, polypeptides, or agonists or antagonists of the invention are used to detect, prevent, diagnose, treat, and/or ameliorate malaria.

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention of the present invention could either be by administering an effective amount of a polypeptide to the patient, or by removing cells from the patient, supplying the cells with a polynucleotide of the present invention, and returning the engineered cells to the patient (ex vivo therapy). Moreover, the polypeptide or polynucleotide of the present invention can be used as an antigen in a vaccine to raise an immune response against infectious disease.

Regeneration

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention can be used to differentiate, proliferate, and attract cells, leading to the regeneration of tissues. (See, Science 276:59-87 (1997)). The regeneration of tissues could be used to repair, replace, or protect tissue damaged by congenital defects, trauma (wounds, burns, incisions, or ulcers), age, disease (e.g. osteoporosis, osteocarthritis, periodontal disease, liver failure), surgery, including cosmetic plastic surgery, fibrosis, reperfusion injury, or systemic cytokine damage.

Tissues that could be regenerated using the present invention include organs (e.g., pancreas, liver, intestine, kidney, skin, endothelium), muscle (smooth, skeletal or cardiac), vasculature (including vascular and lymphatics), nervous, hematopoietic, and skeletal (bone, cartilage, tendon, and ligament) tissue. Preferably, regeneration occurs without or decreased scarring. Regeneration also may include angiogenesis.

Moreover, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may increase regeneration of tissues difficult to heal. For example, increased tendon/ligament regeneration would quicken recovery time after damage. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention could also be used prophylactically in an effort to avoid damage. Specific diseases that could be treated include of tendinitis, carpal tunnel syndrome, and other tendon or ligament defects. A further example of tissue regeneration of non-healing wounds includes pressure ulcers, ulcers associated with vascular insufficiency, surgical, and traumatic wounds.

Similarly, nerve and brain tissue could also be regenerated by using polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, to proliferate and differentiate nerve cells. Diseases that could be treated using this method include central and peripheral nervous system diseases, neuropathies, or mechanical and traumatic disorders (e.g., spinal cord disorders, head trauma, cerebrovascular disease, and stoke). Specifically, diseases associated with peripheral nerve injuries, peripheral neuropathy (e.g., resulting from chemotherapy or other medical therapies), localized neuropathies, and central nervous system diseases (e.g., Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, and Shy-Drager syndrome), could all be treated using the polynucleotides or polypeptides, as well as agonists or antagonists of the present invention.

Gastrointestinal Disorders

Polynucleotides or polypeptides, or agonists or antagonists of the present invention, may be used to detect, prevent, diagnose, prognosticate, treat, and/or ameliorate gastrointestinal diseases and disorders, including inflammatory diseases and/or conditions, infections, cancers (e.g., intestinal neoplasms (carcinoid tumor of the small intestine, non-Hodgkin's lymphoma of the small intestine, small bowl lymphoma)), and ulcers, such as peptic ulcers.

Gastrointestinal disorders include dysphagia, odynophagia, inflammation of the esophagus, peptic esophagitis, gastric reflux, submucosal fibrosis and stricturing, Mallory-Weiss lesions, leiomyomas, lipomas, epidermal cancers, adeoncarcinomas, gastric retention disorders, gastroenteritis, gastric atrophy, gastric/stomach cancers, polyps of the stomach, autoimmune disorders such as pernicious anemia, pyloric stenosis, gastritis (bacterial, viral, eosinophilic, stress-induced, chronic erosive, atrophic, plasma cell, and Menetrier's), and peritoneal diseases (e.g., chyloperioneum, hemoperitoneum, mesenteric cyst, mesenteric lymphadenitis, mesenteric vascular occlusion, panniculitis, neoplasms, peritonitis, pneumoperitoneum, bubphrenic abscess,).

Gastrointestinal disorders also include disorders associated with the small intestine, such as malabsorption syndromes, distension, irritable bowel syndrome, sugar intolerance, celiac disease, duodenal ulcers, duodenitis, tropical sprue, Whipple's disease, intestinal lymphangiectasia, Crohn's disease, appendicitis, obstructions of the ileum, Meckel's diverticulum, multiple diverticula, failure of complete rotation of the small and large intestine, lymphoma, and bacterial and parasitic diseases (such as Traveler's diarrhea, typhoid and paratyphoid, cholera, infection by Roundworms (Ascariasis lumbricoides), Hookworms (Ancylostoma duodenale), Threadworms (Enterobius vermicularis), Tapeworms (Taenia saginata, Echinococcus granulosus, Diphyllobothrium spp., and T solium).

Liver diseases and/or disorders include intrahepatic cholestasis (alagille syndrome, biliary liver cirrhosis), fatty liver (alcoholic fatty liver, reye syndrome), hepatic vein thrombosis, hepatolentricular degeneration, hepatomegaly, hepatopulmonary syndrome, hepatorenal syndrome, portal hypertension (esophageal and gastric varices), liver abscess (amebic liver abscess), liver cirrhosis (alcoholic, biliary and experimental), alcoholic liver diseases (fatty liver, hepatitis, cirrhosis), parasitic (hepatic echinococcosis, fascioliasis, amebic liver abscess), jaundice (hemolytic, hepatocellular, and cholestatic), cholestasis, portal hypertension, liver enlargement, ascites, hepatitis (alcoholic hepatitis, animal hepatitis, chronic hepatitis (autoimmune, hepatitis B, hepatitis C, hepatitis D, drug induced), toxic hepatitis, viral human hepatitis (hepatitis A, hepatitis B, hepatitis C, hepatitis D, hepatitis E), Wilson's disease, granulomatous hepatitis, secondary biliary cirrhosis, hepatic encephalopathy, portal hypertension, varices, hepatic encephalopathy, primary biliary cirrhosis, primary sclerosing cholangitis, hepatocellular adenoma, hemangiomas, bile stones, liver failure (hepatic encephalopathy, acute liver failure), and liver neoplasms (angiomyolipoma, calcified liver metastases, cystic liver metastases, epithelial tumors, fibrolamellar hepatocarcinoma, focal nodular hyperplasia, hepatic adenoma, hepatobiliary cystadenoma, hepatoblastoma, hepatocellular carcinoma, hepatoma, liver cancer, liver hemangioendothelioma, mesenchymal hamartoma, mesenchymal tumors of liver, nodular regenerative hyperplasia, benign liver tumors (Hepatic cysts [Simple cysts, Polycystic liver disease, Hepatobiliary cystadenoma, Choledochal cyst], Mesenchymal tumors [Mesenchymal hamartoma, Infantile hemangioendothelioma, Hemangioma, Peliosis hepatis, Lipomas, Inflammatory pseudotumor, Miscellaneous], Epithelial tumors [Bile duct epithelium (Bile duct hamartoma, Bile duct adenoma), Hepatocyte (Adenoma, Focal nodular hyperplasia, Nodular regenerative hyperplasia)], malignant liver tumors [hepatocellular, hepatoblastoma, hepatocellular carcinoma, cholangiocellular, cholangiocarcinoma, cystadenocarcinoma, tumors of blood vessels, angiosarcoma, Karposi's sarcoma, hemangioendothelioma, other tumors, embryonal sarcoma, fibrosarcoma, leiomyosarcoma, rhabdomyosarcoma, carcinosarcoma, teratoma, carcinoid, squamous carcinoma, primary lymphoma]), peliosis hepatis, erythrohepatic porphyria, hepatic porphyria (acute intermittent porphyria, porphyria cutanea tarda), Zellweger syndrome).

Pancreatic diseases and/or disorders include acute pancreatitis, chronic pancreatitis (acute necrotizing pancreatitis, alcoholic pancreatitis), neoplasms (adenocarcinoma of the pancreas, cystadenocarcinoma, insulinoma, gastrinoma, and glucagonoma, cystic neoplasms, islet-cell tumors, pancreoblastoma), and other pancreatic diseases (e.g., cystic fibrosis, cyst (pancreatic pseudocyst, pancreatic fistula, insufficiency)).

Gallbladder diseases include gallstones (cholelithiasis and choledocholithiasis), postcholecystectomy syndrome, diverticulosis of the gallbladder, acute cholecystitis, chronic cholecystitis, bile duct tumors, and mucocele.

Diseases and/or disorders of the large intestine include antibiotic-associated colitis, diverticulitis, ulcerative colitis, acquired megacolon, abscesses, fungal and bacterial infections, anorectal disorders (e.g., fissures, hemorrhoids), colonic diseases (colitis, colonic neoplasms [colon cancer, adenomatous colon polyps (e.g., villous adenoma), colon carcinoma, colorectal cancer], colonic diverticulitis, colonic diverticulosis, megacolon [Hirschsprung disease, toxic megacolon]; sigmoid diseases [proctocolitis, sigmoin neoplasms]), constipation, Crohn's disease, diarrhea (infantile diarrhea, dysentery), duodenal diseases (duodenal neoplasms, duodenal obstruction, duodenal ulcer, duodenitis), enteritis (enterocolitis), HIV enteropathy, ileal diseases (ileal neoplasms, ileitis), immunoproliferative small intestinal disease, inflammatory bowel disease (ulcerative colitis, Crohn's disease), intestinal atresia, parasitic diseases (anisakiasis, balantidiasis, blastocystis infections, cryptosporidiosis, dientamoebiasis, amebic dysentery, giardiasis), intestinal fistula (rectal fistula), intestinal neoplasms (cecal neoplasms, colonic neoplasms, duodenal neoplasms, ileal neoplasms, intestinal polyps, jejunal neoplasms, rectal neoplasms), intestinal obstruction (afferent loop syndrome, duodenal obstruction, impacted feces, intestinal pseudo-obstruction [cecal volvulus], intussusception), intestinal perforation, intestinal polyps (colonic polyps, gardner syndrome, peutz-jeghers syndrome), jejunal diseases (jejunal neoplasms), malabsorption syndromes (blind loop syndrome, celiac disease, lactose intolerance, short bowl syndrome, tropical sprue, whipple's disease), mesenteric vascular occlusion, pneumatosis cystoides intestinalis, protein-losing enteropathies (intestinal lymphagiectasis), rectal diseases (anus diseases, fecal incontinence, hemorrhoids, proctitis, rectal fistula, rectal prolapse, rectocele), peptic ulcer (duodenal ulcer, peptic esophagitis, hemorrhage, perforation, stomach ulcer, Zollinger-Ellison syndrome), postgastrectomy syndromes (dumping syndrome), stomach diseases (e.g., achlorhydria, duodenogastric reflux (bile reflux), gastric antral vascular ectasia, gastric fistula, gastric outlet obstruction, gastritis (atrophic or hypertrophic), gastroparesis, stomach dilatation, stomach diverticulum, stomach neoplasms (gastric cancer, gastric polyps, gastric adenocarcinoma, hyperplastic gastric polyp), stomach rupture, stomach ulcer, stomach volvulus), tuberculosis, visceroptosis, vomiting (e.g., hematemesis, hyperemesis gravidarum, postoperative nausea and vomiting) and hemorrhagic colitis.

Further diseases and/or disorders of the gastrointestinal system include biliary tract diseases, such as, gastroschisis, fistula (e.g., biliary fistula, esophageal fistula, gastric fistula, intestinal fistula, pancreatic fistula), neoplasms (e.g., biliary tract neoplasms, esophageal neoplasms, such as adenocarcinoma of the esophagus, esophageal squamous cell carcinoma, gastrointestinal neoplasms, pancreatic neoplasms, such as adenocarcinoma of the pancreas, mucinous cystic neoplasm of the pancreas, pancreatic cystic neoplasms, pancreatoblastoma, and peritoneal neoplasms), esophageal disease (e.g., bullous diseases, candidiasis, glycogenic acanthosis, ulceration, barrett esophagus varices, atresia, cyst, diverticulum (e.g., Zenker's diverticulum), fistula (e.g., tracheoesophageal fistula), motility disorders (e.g., CREST syndrome, deglutition disorders, achalasia, spasm, gastroesophageal reflux), neoplasms, perforation (e.g., Boerhaave syndrome, Mallory-Weiss syndrome), stenosis, esophagitis, diaphragmatic hernia (e.g., hiatal hernia); gastrointestinal diseases, such as, gastroenteritis (e.g., cholera morbus, norwalk virus infection), hemorrhage (e.g., hematemesis, melena, peptic ulcer hemorrhage), stomach neoplasms (gastric cancer, gastric polyps, gastric adenocarcinoma, stomach cancer)), hernia (e.g., congenital diaphragmatic hernia, femoral hernia, inguinal hernia, obturator hernia, umbilical hernia, ventral hernia), and intestinal diseases (e.g., cecal diseases (appendicitis, cecal neoplasms)).

Chemotaxis

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may have chemotaxis activity. A chemotaxic molecule attracts or mobilizes cells (e.g., monocytes, fibroblasts, neutrophils, T-cells, mast cells, eosinophils, epithelial and/or endothelial cells) to a particular site in the body, such as inflammation, infection, or site of hyperproliferation. The mobilized cells can then fight off and/or heal the particular trauma or abnormality.

Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may increase chemotaxic activity of particular cells. These chemotactic molecules can then be used to treat inflammation, infection, hyperproliferative disorders, or any immune system disorder by increasing the number of cells targeted to a particular location in the body. For example, chemotaxic molecules can be used to treat wounds and other trauma to tissues by attracting immune cells to the injured location. Chemotactic molecules of the present invention can also attract fibroblasts, which can be used to treat wounds.

It is also contemplated that polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may inhibit chemotactic activity. These molecules could also be used to treat disorders. Thus, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention could be used as an inhibitor of chemotaxis.

Binding Activity

A polypeptide of the present invention may be used to screen for molecules that bind to the polypeptide or for molecules to which the polypeptide binds. The binding of the polypeptide and the molecule may activate (agonist), increase, inhibit (antagonist), or decrease activity of the polypeptide or the molecule bound. Examples of such molecules include antibodies, oligonucleotides, proteins (e.g., receptors), or small molecules.

Preferably, the molecule is closely related to the natural ligand of the polypeptide, e.g., a fragment of the ligand, or a natural substrate, a ligand, a structural or functional mimetic. (See, Coligan et al., Current Protocols in Immunology 1(2):Chapter 5 (1991)). Similarly, the molecule can be closely related to the natural receptor to which the polypeptide binds, or at least, a fragment of the receptor capable of being bound by the polypeptide (e.g., active site). In either case, the molecule can be rationally designed using known techniques.

Preferably, the screening for these molecules involves producing appropriate cells which express the polypeptide. Preferred cells include cells from mammals, yeast, Drosophila, or E. coli. Cells expressing the polypeptide (or cell membrane containing the expressed polypeptide) are then preferably contacted with a test compound potentially containing the molecule to observe binding, stimulation, or inhibition of activity of either the polypeptide or the molecule.

The assay may simply test binding of a candidate compound to the polypeptide, wherein binding is detected by a label, or in an assay involving competition with a labeled competitor. Further, the assay may test whether the candidate compound results in a signal generated by binding to the polypeptide.

Alternatively, the assay can be carried out using cell-free preparations, polypeptide/molecule affixed to a solid support, chemical libraries, or natural product mixtures. The assay may also simply comprise the steps of mixing a candidate compound with a solution containing a polypeptide, measuring polypeptide/molecule activity or binding, and comparing the polypeptide/molecule activity or binding to a standard.

Preferably, an ELISA assay can measure polypeptide level or activity in a sample (e.g., biological sample) using a monoclonal or polyclonal antibody. The antibody can measure polypeptide level or activity by either binding, directly or indirectly, to the polypeptide or by competing with the polypeptide for a substrate.

Additionally, the receptor to which the polypeptide of the present invention binds can be identified by numerous methods known to those of skill in the art, for example, ligand panning and FACS sorting (Coligan, et al., Current Protocols in Immun., 1(2), Chapter 5, (1991)). For example, expression cloning is employed wherein polyadenylated RNA is prepared from a cell responsive to the polypeptides, for example, NIH3T3 cells which are known to contain multiple receptors for the FGF family proteins, and SC-3 cells, and a cDNA library created from this RNA is divided into pools and used to transfect COS cells or other cells that are not responsive to the polypeptides. Transfected cells which are grown on glass slides are exposed to the polypeptide of the present invention, after they have been labeled. The polypeptides can be labeled by a variety of means including iodination or inclusion of a recognition site for a site-specific protein kinase.

Following fixation and incubation, the slides are subjected to auto-radiographic analysis. Positive pools are identified and sub-pools are prepared and re-transfected using an iterative sub-pooling and re-screening process, eventually yielding a single clones that encodes the putative receptor.

As an alternative approach for receptor identification, the labeled polypeptides can be photoaffinity linked with cell membrane or extract preparations that express the receptor molecule. Cross-linked material is resolved by PAGE analysis and exposed to X-ray film. The labeled complex containing the receptors of the polypeptides can be excised, resolved into peptide fragments, and subjected to protein microsequencing. The amino acid sequence obtained from microsequencing would be used to design a set of degenerate oligonucleotide probes to screen a cDNA library to identify the genes encoding the putative receptors.

Moreover, the techniques of gene-shuffling, motif-shuffling, exon-shuffling, and/or codon-shuffling (collectively referred to as “DNA shuffling”) may be employed to modulate the activities of the polypeptide of the present invention thereby effectively generating agonists and antagonists of the polypeptide of the present invention. See generally, U.S. Pat. Nos. 5,605,793, 5,811,238, 5,830,721, 5,834,252, and 5,837,458, and Patten, P. A., et al., Curr. Opinion Biotechnol. 8:724-33 (1997); Harayama, S. Trends Biotechnol. 16(2):76-82 (1998); Hansson, L. O., et al., J. Mol. Biol. 287:265-76 (1999); and Lorenzo, M. M. and Blasco, R. Biotechniques 24(2):308-13 (1998); each of these patents and publications are hereby incorporated by reference). In one embodiment, alteration of polynucleotides and corresponding polypeptides may be achieved by DNA shuffling. DNA shuffling involves the assembly of two or more DNA segments into a desired molecule by homologous, or site-specific, recombination. In another embodiment, polynucleotides and corresponding polypeptides may be altered by being subjected to random mutagenesis by error-prone PCR, random nucleotide insertion or other methods prior to recombination. In another embodiment, one or more components, motifs, sections, parts, domains, fragments, etc., of the polypeptide of the present invention may be recombined with one or more components, motifs, sections, parts, domains, fragments, etc. of one or more heterologous molecules. In preferred embodiments, the heterologous molecules are family members. In further preferred embodiments, the heterologous molecule is a growth factor such as, for example, platelet-derived growth factor (PDGF), insulin-like growth factor (IGF-I), transforming growth factor (TGF)-alpha, epidermal growth factor (EGF), fibroblast growth factor (FGF), TGF-beta, bone morphogenetic protein (BMP)-2, BMP-4, BMP-5, BMP-6, BMP-7, activins A and B, decapentaplegic(dpp), 60A, OP-2, dorsalin, growth differentiation factors (GDFs), nodal, MIS, inhibin-alpha, TGF-beta1, TGF-beta2, TGF-beta3, TGF-beta5, and glial-derived neurotrophic factor (GDNF).

Other preferred fragments are biologically active fragments of the polypeptide of the present invention. Biologically active fragments are those exhibiting activity similar, but not necessarily identical, to an activity of the polypeptide of the present invention. The biological activity of the fragments may include an improved desired activity, or a decreased undesirable activity.

Additionally, this invention provides a method of screening compounds to identify those which modulate the action of the polypeptide of the present invention. An example of such an assay comprises combining a mammalian fibroblast cell, a the polypeptide of the present invention, the compound to be screened and ³[H] thymidine under cell culture conditions where the fibroblast cell would normally proliferate. A control assay may be performed in the absence of the compound to be screened and compared to the amount of fibroblast proliferation in the presence of the compound to determine if the compound stimulates proliferation by determining the uptake of ³[H] thymidine in each case. The amount of fibroblast cell proliferation is measured by liquid scintillation chromatography which measures the incorporation of ³[H] thymidine. Both agonist and antagonist compounds may be identified by this procedure.

In another method, a mammalian cell or membrane preparation expressing a receptor for a polypeptide of the present invention is incubated with a labeled polypeptide of the present invention in the presence of the compound. The ability of the compound to enhance or block this interaction could then be measured. Alternatively, the response of a known second messenger system following interaction of a compound to be screened and the receptor is measured and the ability of the compound to bind to the receptor and elicit a second messenger response is measured to determine if the compound is a potential agonist or antagonist. Such second messenger systems include but are not limited to, cAMP guanylate cyclase, ion channels or phosphoinositide hydrolysis.

All of these above assays can be used as diagnostic or prognostic markers. The molecules discovered using these assays can be used to treat disease or to bring about a particular result in a patient (e.g., blood vessel growth) by activating or inhibiting the polypeptide/molecule. Moreover, the assays can discover agents which may inhibit or enhance the production of the polypeptides of the invention from suitably manipulated cells or tissues.

Therefore, the invention includes a method of identifying compounds which bind to a polypeptide of the invention comprising the steps of: (a) incubating a candidate binding compound with a polypeptide of the present invention; and (b) determining if binding has occurred. Moreover, the invention includes a method of identifying agonists/antagonists comprising the steps of: (a) incubating a candidate compound with a polypeptide of the present invention, (b) assaying a biological activity, and (b) determining if a biological activity of the polypeptide has been altered.

Targeted Delivery

In another embodiment, the invention provides a method of delivering compositions to targeted cells expressing a receptor for a polypeptide of the invention, or cells expressing a cell bound form of a polypeptide of the invention.

As discussed herein, polypeptides or antibodies of the invention may be associated with heterologous polypeptides, heterologous nucleic acids, toxins, or prodrugs via hydrophobic, hydrophilic, ionic and/or covalent interactions. In one embodiment, the invention provides a method for the specific delivery of compositions of the invention to cells by administering polypeptides of the invention (including antibodies) that are associated with heterologous polypeptides or nucleic acids. In one example, the invention provides a method for delivering a therapeutic protein into the targeted cell. In another example, the invention provides a method for delivering a single stranded nucleic acid (e.g., antisense or ribozymes) or double stranded nucleic acid (e.g., DNA that can integrate into the cell's genome or replicate episomally and that can be transcribed) into the targeted cell.

In another embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention (e.g., polypeptides of the invention or antibodies of the invention) in association with toxins or cytotoxic prodrugs.

By “toxin” is meant compounds that bind and activate endogenous cytotoxic effector systems, radioisotopes, holotoxins, modified toxins, catalytic subunits of toxins, or any molecules or enzymes not normally present in or on the surface of a cell that under defined conditions cause the cell's death. Toxins that may be used according to the methods of the invention include, but are not limited to, radioisotopes known in the art, compounds such as, for example, antibodies (or complement fixing containing portions thereof) that bind an inherent or induced endogenous cytotoxic effector system, thymidine kinase, endonuclease, RNAse, alpha toxin, ricin, abrin, Pseudomonas exotoxin A, diphtheria toxin, saporin, momordin, gelonin, pokeweed antiviral protein, alpha-sarcin and cholera toxin. By “cytotoxic prodrug” is meant a non-toxic compound that is converted by an enzyme, normally present in the cell, into a cytotoxic compound. Cytotoxic prodrugs that may be used according to the methods of the invention include, but are not limited to, glutamyl derivatives of benzoic acid mustard alkylating agent, phosphate derivatives of etoposide or mitomycin C, cytosine arabinoside, daunorubisin, and phenoxyacetamide derivatives of doxorubicin.

Drug Screening

Further contemplated is the use of the polypeptides of the present invention, or the polynucleotides encoding these polypeptides, to screen for molecules which modify the activities of the polypeptides of the present invention. Such a method would include contacting the polypeptide of the present invention with a selected compound(s) suspected of having antagonist or agonist activity, and assaying the activity of these polypeptides following binding.

This invention is particularly useful for screening therapeutic compounds by using the polypeptides of the present invention, or binding fragments thereof, in any of a variety of drug screening techniques. The polypeptide or fragment employed in such a test may be affixed to a solid support, expressed on a cell surface, free in solution, or located intracellularly. One method of drug screening utilizes eukaryotic or prokaryotic host cells which are stably transformed with recombinant nucleic acids expressing the polypeptide or fragment. Drugs are screened against such transformed cells in competitive binding assays. One may measure, for example, the formulation of complexes between the agent being tested and a polypeptide of the present invention.

Thus, the present invention provides methods of screening for drugs or any other agents which affect activities mediated by the polypeptides of the present invention. These methods comprise contacting such an agent with a polypeptide of the present invention or a fragment thereof and assaying for the presence of a complex between the agent and the polypeptide or a fragment thereof, by methods well known in the art. In such a competitive binding assay, the agents to screen are typically labeled. Following incubation, free agent is separated from that present in bound form, and the amount of free or uncomplexed label is a measure of the ability of a particular agent to bind to the polypeptides of the present invention.

Another technique for drug screening provides high throughput screening for compounds having suitable binding affinity to the polypeptides of the present invention, and is described in great detail in European Patent Application 84/03564, published on Sep. 13, 1984, which is incorporated herein by reference herein. Briefly stated, large numbers of different small peptide test compounds are synthesized on a solid substrate, such as plastic pins or some other surface. The peptide test compounds are reacted with polypeptides of the present invention and washed. Bound polypeptides are then detected by methods well known in the art. Purified polypeptides are coated directly onto plates for use in the aforementioned drug screening techniques. In addition, non-neutralizing antibodies may be used to capture the peptide and immobilize it on the solid support.

This invention also contemplates the use of competitive drug screening assays in which neutralizing antibodies capable of binding polypeptides of the present invention specifically compete with a test compound for binding to the polypeptides or fragments thereof. In this manner, the antibodies are used to detect the presence of any peptide which shares one or more antigenic epitopes with a polypeptide of the invention.

Antisense and Ribozyme (Antagonists)

In specific embodiments, antagonists according to the present invention are nucleic acids corresponding to the sequences contained in SEQ ID NO:X, or the complementary strand thereof, and/or to cDNA sequences contained in cDNA ATCC™ Deposit No:Z identified for example, in Table 1A and/or 1B. In one embodiment, antisense sequence is generated internally, by the organism, in another embodiment, the antisense sequence is separately administered (see, for example, O'Connor, J., Neurochem. 56:560 (1991). Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988). Antisense technology can be used to control gene expression through antisense DNA or RNA, or through triple-helix formation. Antisense techniques are discussed for example, in Okano, J., Neurochem. 56:560 (1991); Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988). Triple helix formation is discussed in, for instance, Lee et al., Nucleic Acids Research 6:3073 (1979); Cooney et al., Science 241:456 (1988); and Dervan et al., Science 251:1300 (1991). The methods are based on binding of a polynucleotide to a complementary DNA or RNA.

For example, the use of c-myc and c-myb antisense RNA constructs to inhibit the growth of the non-lymphocytic leukemia cell line HL-60 and other cell lines was previously described. (Wickstrom et al. (1988); Anfossi et al. (1989)). These experiments were performed in vitro by incubating cells with the oligoribonucleotide. A similar procedure for in vivo use is described in WO 91/15580. Briefly, a pair of oligonucleotides for a given antisense RNA is produced as follows: A sequence complimentary to the first 15 bases of the open reading frame is flanked by an EcoRI site on the 5 end and a HindIII site on the 3 end. Next, the pair of oligonucleotides is heated at 90° C. for one minute and then annealed in 2× ligation buffer (20 mM TRIS HCl pH 7.5, 10 mM MgCl₂, 10mM dithiothreitol (DTT) and 0.2 mM ATP) and then ligated to the EcoRI/Hind III site of the retroviral vector PMV7 (WO 91/15580).

For example, the 5′ coding portion of a polynucleotide that encodes the polypeptide of the present invention may be used to design an antisense RNA oligonucleotide of from about 10 to 40 base pairs in length. A DNA oligonucleotide is designed to be complementary to a region of the gene involved in transcription thereby preventing transcription and the production of the receptor. The antisense RNA oligonucleotide hybridizes to the mRNA in vivo and blocks translation of the mRNA molecule into receptor polypeptide.

In one embodiment, the antisense nucleic acid of the invention is produced intracellularly by transcription from an exogenous sequence. For example, a vector or a portion thereof, is transcribed, producing an antisense nucleic acid (RNA) of the invention. Such a vector would contain a sequence encoding the antisense nucleic acid. Such a vector can remain episomal or become chromosomally integrated, as long as it can be transcribed to produce the desired antisense RNA. Such vectors can be constructed by recombinant DNA technology methods standard in the art. Vectors can be plasmid, viral, or others known in the art, used for replication and expression in vertebrate cells. Expression of the sequence encoding the polypeptide of the present invention or fragments thereof, can be by any promoter known in the art to act in vertebrate, preferably human cells. Such promoters can be inducible or constitutive. Such promoters include, but are not limited to, the SV40 early promoter region (Bernoist and Chambon, Nature 29:304-310 (1981), the promoter contained in the 3′ long terminal repeat of Rous sarcoma virus (Yamamoto et al., Cell 22:787-797 (1980), the herpes thymidine promoter (Wagner et al., Proc. Natl. Acad. Sci. U.S.A. 78:1441-1445 (1981), the regulatory sequences of the metallothionein gene (Brinster, et al., Nature 296:39-42 (1982)), etc.

The antisense nucleic acids of the invention comprise a sequence complementary to at least a portion of an RNA transcript of a gene of the present invention. However, absolute complementarity, although preferred, is not required. A sequence “complementary to at least a portion of an RNA,” referred to herein, means a sequence having sufficient complementarity to be able to hybridize with the RNA, forming a stable duplex; in the case of double stranded antisense nucleic acids, a single strand of the duplex DNA may thus be tested, or triplex formation may be assayed. The ability to hybridize will depend on both the degree of complementarity and the length of the antisense nucleic acid. Generally, the larger the hybridizing nucleic acid, the more base mismatches with a RNA it may contain and still form a stable duplex (or triplex as the case may be). One skilled in the art can ascertain a tolerable degree of mismatch by use of standard procedures to determine the melting point of the hybridized complex.

Oligonucleotides that are complementary to the 5′ end of the message, e.g., the 5′ untranslated sequence up to and including the AUG initiation codon, should work most efficiently at inhibiting translation. However, sequences complementary to the 3′ untranslated sequences of mRNAs have been shown to be effective at inhibiting translation of mRNAs as well. See generally, Wagner, R., 1994, Nature 372:333-335. Thus, oligonucleotides complementary to either the 5′- or 3′-non-translated, non-coding regions of polynucleotide sequences described herein could be used in an antisense approach to inhibit translation of endogenous mRNA. Oligonucleotides complementary to the 5′ untranslated region of the mRNA should include the complement of the AUG start codon. Antisense oligonucleotides complementary to mRNA coding regions are less efficient inhibitors of translation but could be used in accordance with the invention. Whether designed to hybridize to the 5′-, 3′- or coding region of mRNA of the present invention, antisense nucleic acids should be at least six nucleotides in length, and are preferably oligonucleotides ranging from 6 to about 50 nucleotides in length. In specific aspects the oligonucleotide is at least 10 nucleotides, at least 17 nucleotides, at least 25 nucleotides or at least 50 nucleotides.

The polynucleotides of the invention can be DNA or RNA or chimeric mixtures or derivatives or modified versions thereof, single-stranded or double-stranded. The oligonucleotide can be modified at the base moiety, sugar moiety, or phosphate backbone, for example, to improve stability of the molecule, hybridization, etc. The oligonucleotide may include other appended groups such as peptides (e.g., for targeting host cell receptors in vivo), or agents facilitating transport across the cell membrane (see, e.g., Letsinger et al., 1989, Proc. Natl. Acad. Sci. U.S.A. 86:6553-6556; Lemaitre et al., 1987, Proc. Natl. Acad. Sci. 84:648-652; PCT Publication No. WO88/09810, published Dec. 15, 1988) or the blood-brain barrier (see, e.g., PCT Publication No. WO89/10134, published Apr. 25, 1988), hybridization-triggered cleavage agents. (See, e.g., Krol et al., 1988, BioTechniques 6:958-976) or intercalating agents. (See, e.g., Zon, 1988, Pharm. Res. 5:539-549). To this end, the oligonucleotide may be conjugated to another molecule, e.g., a peptide, hybridization triggered cross-linking agent, transport agent, hybridization-triggered cleavage agent, etc.

The antisense oligonucleotide may comprise at least one modified base moiety which is selected from the group including, but not limited to, 5-fluorouracil, 5-bromouracil, 5-chlorouracil, 5-iodouracil, hypoxanthine, xantine, 4-acetylcytosine, 5-(carboxyhydroxylmethyl) uracil, 5-carboxymethylaminomethyl-2-thiouridine, 5-carboxymethylaminomethyluracil, dihydrouracil, beta-D-galactosylqueosine, inosine, N6-isopentenyladenine, 1-methylguanine, 1-methylinosine, 2,2-dimethylguanine, 2-methyladenine, 2-methylguanine, 3-methylcytosine, 5-methylcytosine, N6-adenine, 7-methylguanine, 5-methylaminomethyluracil, 5-methoxyaminomethyl-2-thiouracil, beta-D-mannosylqueosine, 5′-methoxycarboxymethyluracil, 5-methoxyuracil, 2-methylthio-N-6-isopentenyladenine, uracil-5-oxyacetic acid (v), wybutoxosine, pseudouracil, queosine, 2-thiocytosine, 5-methyl-2-thiouracil, 2-thiouracil, 4-thiouracil, 5-methyluracil, uracil-5-oxyacetic acid methylester, uracil-5-oxyacetic acid (v), 5-methyl-2-thiouracil, 3-(3-amino-3-N-2-carboxypropyl) uracil, (acp3)w, and 2,6-diaminopurine.

The antisense oligonucleotide may also comprise at least one modified sugar moiety selected from the group including, but not limited to, arabinose, 2-fluoroarabinose, xylulose, and hexose.

In yet another embodiment, the antisense oligonucleotide comprises at least one modified phosphate backbone selected from the group including, but not limited to, a phosphorothioate, a phosphorodithioate, a phosphoramidothioate, a phosphoramidate, a phosphordiamidate, a methylphosphonate, an alkyl phosphotriester, and a formacetal or analog thereof.

In yet another embodiment, the antisense oligonucleotide is an a-anomeric oligonucleotide. An a-anomeric oligonucleotide forms specific double-stranded hybrids with complementary RNA in which, contrary to the usual b-units, the strands run parallel to each other (Gautier et al., 1987, Nucl. Acids Res. 15:6625-6641). The oligonucleotide is a 2′-O-methylribonucleotide (Inoue et al., 1987, Nucl. Acids Res. 15:6131-6148), or a chimeric RNA-DNA analogue (Inoue et al., 1987, FEBS Lett. 215:327-330).

Polynucleotides of the invention may be synthesized by standard methods known in the art, e.g. by use of an automated DNA synthesizer (such as are commercially available from Biosearch, Applied Biosystems, etc.). As examples, phosphorothioate oligonucleotides may be synthesized by the method of Stein et al. (1988, Nucl. Acids Res. 16:3209), methylphosphonate oligonucleotides can be prepared by use of controlled pore glass polymer supports (Sarin et al., 1988, Proc. Natl. Acad. Sci. U.S.A. 85:7448-7451), etc.

While antisense nucleotides complementary to the coding region sequence could be used, those complementary to the transcribed untranslated region are most preferred.

Potential antagonists according to the invention also include catalytic RNA, or a ribozyme (See, e.g., PCT International Publication WO 90/11364, published Oct. 4, 1990; Sarver et al, Science 247:1222-1225 (1990). While ribozymes that cleave mRNA at site specific recognition sequences can be used to destroy mRNAs, the use of hammerhead ribozymes is preferred. Hammerhead ribozymes cleave mRNAs at locations dictated by flanking regions that form complementary base pairs with the target mRNA. The sole requirement is that the target mRNA have the following sequence of two bases: 5′-UG-3′. The construction and production of hammerhead ribozymes is well known in the art and is described more fully in Haseloff and Gerlach, Nature 334:585-591 (1988). There are numerous potential hammerhead ribozyme cleavage sites within the nucleotide sequence of SEQ ID NO:X. Preferably, the ribozyme is engineered so that the cleavage recognition site is located near the 5′ end of the mRNA; i.e., to increase efficiency and minimize the intracellular accumulation of non-functional mRNA transcripts.

As in the antisense approach, the ribozymes of the invention can be composed of modified oligonucleotides (e.g., for improved stability, targeting, etc.) and should be delivered to cells which express in vivo. DNA constructs encoding the ribozyme may be introduced into the cell in the same manner as described above for the introduction of antisense encoding DNA. A preferred method of delivery involves using a DNA construct “encoding” the ribozyme under the control of a strong constitutive promoter, such as, for example, pol III or pol II promoter, so that transfected cells will produce sufficient quantities of the ribozyme to destroy endogenous messages and inhibit translation. Since ribozymes unlike antisense molecules, are catalytic, a lower intracellular concentration is required for efficiency.

Antagonist/agonist compounds may be employed to inhibit the cell growth and proliferation effects of the polypeptides of the present invention on neoplastic cells and tissues, i.e. stimulation of angiogenesis of tumors, and, therefore, retard or prevent abnormal cellular growth and proliferation, for example, in tumor formation or growth.

The antagonist/agonist may also be employed to prevent hyper-vascular diseases, and prevent the proliferation of epithelial lens cells after extracapsular cataract surgery. Prevention of the mitogenic activity of the polypeptides of the present invention may also be desirous in cases such as restenosis after balloon angioplasty.

The antagonist/agonist may also be employed to prevent the growth of scar tissue during wound healing.

The antagonist/agonist may also be employed to treat the diseases described herein.

Thus, the invention provides a method of treating disorders or diseases, including but not limited to the disorders or diseases listed throughout this application, associated with overexpression of a polynucleotide of the present invention by administering to a patient (a) an antisense molecule directed to the polynucleotide of the present invention, and/or (b) a ribozyme directed to the polynucleotide of the present invention.

Binding Peptides and Other Molecules

The invention also encompasses screening methods for identifying polypeptides and nonpolypeptides that bind polypeptides of the invention, and the binding molecules identified thereby. These binding molecules are useful, for example, as agonists and antagonists of the polypeptides of the invention. Such agonists and antagonists can be used, in accordance with the invention, in the therapeutic embodiments described in detail, below.

This method comprises the steps of:

a. contacting polypeptides of the invention with a plurality of molecules; and

b. identifying a molecule that binds the polypeptides of the invention.

The step of contacting the polypeptides of the invention with the plurality of molecules may be effected in a number of ways. For example, one may contemplate immobilizing the polypeptides on a solid support and bringing a solution of the plurality of molecules in contact with the immobilized polypeptides. Such a procedure would be akin to an affinity chromatographic process, with the affinity matrix being comprised of the immobilized polypeptides of the invention. The molecules having a selective affinity for the polypeptides can then be purified by affinity selection. The nature of the solid support, process for attachment of the polypeptides to the solid support, solvent, and conditions of the affinity isolation or selection are largely conventional and well known to those of ordinary skill in the art.

Alternatively, one may also separate a plurality of polypeptides into substantially separate fractions comprising a subset of or individual polypeptides. For instance, one can separate the plurality of polypeptides by gel electrophoresis, column chromatography, or like method known to those of ordinary skill for the separation of polypeptides. The individual polypeptides can also be produced by a transformed host cell in such a way as to be expressed on or about its outer surface (e.g., a recombinant phage). Individual isolates can then be “probed” by the polypeptides of the invention, optionally in the presence of an inducer should one be required for expression, to determine if any selective affinity interaction takes place between the polypeptides and the individual clone. Prior to contacting the polypeptides with each fraction comprising individual polypeptides, the polypeptides could first be transferred to a solid support for additional convenience. Such a solid support may simply be a piece of filter membrane, such as one made of nitrocellulose or nylon. In this manner, positive clones could be identified from a collection of transformed host cells of an expression library, which harbor a DNA construct encoding a polypeptide having a selective affinity for polypeptides of the invention. Furthermore, the amino acid sequence of the polypeptide having a selective affinity for the polypeptides of the invention can be determined directly by conventional means or the coding sequence of the DNA encoding the polypeptide can frequently be determined more conveniently. The primary sequence can then be deduced from the corresponding DNA sequence. If the amino acid sequence is to be determined from the polypeptide itself, one may use microsequencing techniques. The sequencing technique may include mass spectroscopy.

In certain situations, it may be desirable to wash away any unbound polypeptides from a mixture of the polypeptides of the invention and the plurality of polypeptides prior to attempting to determine or to detect the presence of a selective affinity interaction. Such a wash step may be particularly desirable when the polypeptides of the invention or the plurality of polypeptides are bound to a solid support.

The plurality of molecules provided according to this method may be provided by way of diversity libraries, such as random or combinatorial peptide or nonpeptide libraries which can be screened for molecules that specifically bind polypeptides of the invention. Many libraries are known in the art that can be used, e.g., chemically synthesized libraries, recombinant (e.g., phage display libraries), and in vitro translation-based libraries. Examples of chemically synthesized libraries are described in Fodor et al., 1991, Science 251:767-773; Houghten et al., 1991, Nature 354:84-86; Lam et al., 1991, Nature 354:82-84; Medynski, 1994, Bio/Technology 12:709-710; Gallop et al., 1994, J. Medicinal Chemistry 37(9):1233-1251; Ohlmeyer et al., 1993, Proc. Natl. Acad. Sci. USA 90:10922-10926; Erb et al., 1994, Proc. Natl. Acad. Sci. USA 91:11422-11426; Houghten et al., 1992, Biotechniques 13:412; Jayawickreme et al., 1994, Proc. Natl. Acad. Sci. USA 91:1614-1618; Salmon et al., 1993, Proc. Natl. Acad. Sci. USA 90:11708-11712; PCT Publication No. WO 93/20242; and Brenner and Lerner, 1992, Proc. Natl. Acad. Sci. USA 89:5381-5383.

Examples of phage display libraries are described in Scott and Smith, 1990, Science 249:386-390; Devlin et al., 1990, Science, 249:404-406; Christian, R. B., et al., 1992, J. Mol. Biol. 227:711-718); Lenstra, 1992, J. Immunol. Meth. 152:149-157; Kay et al., 1993, Gene 128:59-65; and PCT Publication No. WO 94/18318 dated Aug. 18, 1994.

In vitro translation-based libraries include but are not limited to those described in PCT Publication No. WO 91/05058 dated Apr. 18, 1991; and Mattheakis et al., 1994, Proc. Natl. Acad. Sci. USA 91:9022-9026.

By way of examples of nonpeptide libraries, a benzodiazepine library (see e.g., Bunin et al., 1994, Proc. Natl. Acad. Sci. USA 91:4708-4712) can be adapted for use. Peptoid libraries (Simon et al., 1992, Proc. Natl. Acad. Sci. USA 89:9367-9371) can also be used. Another example of a library that can be used, in which the amide functionalities in peptides have been permethylated to generate a chemically transformed combinatorial library, is described by Ostresh et al. (1994, Proc. Natl. Acad. Sci. USA 91:11138-11142).

The variety of non-peptide libraries that are useful in the present invention is great. For example, Ecker and Crooke, 1995, Bio/Technology 13:351-360 list benzodiazepines, hydantoins, piperazinediones, biphenyls, sugar analogs, beta-mercaptoketones, arylacetic acids, acylpiperidines, benzopyrans, cubanes, xanthines, aminimides, and oxazolones as among the chemical species that form the basis of various libraries.

Non-peptide libraries can be classified broadly into two types: decorated monomers and oligomers. Decorated monomer libraries employ a relatively simple scaffold structure upon which a variety functional groups is added. Often the scaffold will be a molecule with a known useful pharmacological activity. For example, the scaffold might be the benzodiazepine structure.

Non-peptide oligomer libraries utilize a large number of monomers that are assembled together in ways that create new shapes that depend on the order of the monomers. Among the monomer units that have been used are carbamates, pyrrolinones, and morpholinos. Peptoids, peptide-like oligomers in which the side chain is attached to the alpha amino group rather than the alpha carbon, form the basis of another version of non-peptide oligomer libraries. The first non-peptide oligomer libraries utilized a single type of monomer and thus contained a repeating backbone. Recent libraries have utilized more than one monomer, giving the libraries added flexibility.

Screening the libraries can be accomplished by any of a variety of commonly known methods. See, e.g., the following references, which disclose screening of peptide libraries: Parmley and Smith, 1989, Adv. Exp. Med. Biol. 251:215-218; Scott and Smith, 1990, Science 249:386-390; Fowlkes et al., 1992; BioTechniques 13:422-427; Oldenburg et al., 1992, Proc. Natl. Acad. Sci. USA 89:5393-5397; Yu et al., 1994, Cell 76:933-945; Staudt et al., 1988, Science 241:577-580; Bock et al., 1992, Nature 355:564-566; Tuerk et al., 1992, Proc. Natl. Acad. Sci. USA 89:6988-6992; Ellington et al., 1992, Nature 355:850-852; U.S. Pat. No. 5,096,815, U.S. Pat. No. 5,223,409, and U.S. Pat. No. 5,198,346, all to Ladner et al.; Rebar and Pabo, 1993, Science 263:671-673; and CT Publication No. WO 94/18318.

In a specific embodiment, screening to identify a molecule that binds polypeptides of the invention can be carried out by contacting the library members with polypeptides of the invention immobilized on a solid phase and harvesting those library members that bind to the polypeptides of the invention. Examples of such screening methods, termed “panning” techniques are described by way of example in Parmley and Smith, 1988, Gene 73:305-318; Fowlkes et al., 1992, BioTechniques 13:422-427; PCT Publication No. WO 94/18318; and in references cited herein.

In another embodiment, the two-hybrid system for selecting interacting proteins in yeast (Fields and Song, 1989, Nature 340:245-246; Chien et al., 1991, Proc. Natl. Acad. Sci. USA 88:9578-9582) can be used to identify molecules that specifically bind to polypeptides of the invention.

Where the binding molecule is a polypeptide, the polypeptide can be conveniently selected from any peptide library, including random peptide libraries, combinatorial peptide libraries, or biased peptide libraries. The term “biased” is used herein to mean that the method of generating the library is manipulated so as to restrict one or more parameters that govern the diversity of the resulting collection of molecules, in this case peptides.

Thus, a truly random peptide library would generate a collection of peptides in which the probability of finding a particular amino acid at a given position of the peptide is the same for all 20 amino acids. A bias can be introduced into the library, however, by specifying, for example, that a lysine occur every fifth amino acid or that positions 4, 8, and 9 of a decapeptide library be fixed to include only arginine. Clearly, many types of biases can be contemplated, and the present invention is not restricted to any particular bias. Furthermore, the present invention contemplates specific types of peptide libraries, such as phage displayed peptide libraries and those that utilize a DNA construct comprising a lambda phage vector with a DNA insert.

As mentioned above, in the case of a binding molecule that is a polypeptide, the polypeptide may have about 6 to less than about 60 amino acid residues, preferably about 6 to about 10 amino acid residues, and most preferably, about 6 to about 22 amino acids. In another embodiment, a binding polypeptide has in the range of 15-100 amino acids, or 20-50 amino acids.

The selected binding polypeptide can be obtained by chemical synthesis or recombinant expression.

Other Activities

A polypeptide, polynucleotide, agonist, or antagonist of the present invention, as a result of the ability to stimulate vascular endothelial cell growth, may be employed in treatment for stimulating re-vascularization of ischemic tissues due to various disease conditions such as thrombosis, arteriosclerosis, and other cardiovascular conditions. The polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed to stimulate angiogenesis and limb regeneration, as discussed above.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed for treating wounds due to injuries, burns, post-operative tissue repair, and ulcers since they are mitogenic to various cells of different origins, such as fibroblast cells and skeletal muscle cells, and therefore, facilitate the repair or replacement of damaged or diseased tissue.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed stimulate neuronal growth and to treat and prevent neuronal damage which occurs in certain neuronal disorders or neuro-degenerative conditions such as Alzheimer's disease, Parkinson's disease, and AIDS-related complex. A polypeptide, polynucleotide, agonist, or antagonist of the present invention may have the ability to stimulate chondrocyte growth, therefore, they may be employed to enhance bone and periodontal regeneration and aid in tissue transplants or bone grafts.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may be also be employed to prevent skin aging due to sunburn by stimulating keratinocyte growth.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed for preventing hair loss, since FGF family members activate hair-forming cells and promotes melanocyte growth. Along the same lines, a polypeptide, polynucleotide, agonist, or antagonist of the present invention may be employed to stimulate growth and differentiation of hematopoietic cells and bone marrow cells when used in combination with other cytokines.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed to maintain organs before transplantation or for supporting cell culture of primary tissues. A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed for inducing tissue of mesodermal origin to differentiate in early embryos.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also increase or decrease the differentiation or proliferation of embryonic stem cells, besides, as discussed above, hematopoietic lineage.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be used to modulate mammalian characteristics, such as body height, weight, hair color, eye color, skin, percentage of adipose tissue, pigmentation, size, and shape (e.g., cosmetic surgery). Similarly, a polypeptide, polynucleotide, agonist, or antagonist of the present invention may be used to modulate mammalian metabolism affecting catabolism, anabolism, processing, utilization, and storage of energy.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may be used to change a mammal's mental state or physical state by influencing biorhythms, caricadic rhythms, depression (including depressive disorders), tendency for violence, tolerance for pain, reproductive capabilities (preferably by Activin or Inhibin-like activity), hormonal or endocrine levels, appetite, libido, memory, stress, or other cognitive qualities.

A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be used as a food additive or preservative, such as to increase or decrease storage capabilities, fat content, lipid, protein, carbohydrate, vitamins, minerals, cofactors or other nutritional components.

The above-recited applications have uses in a wide variety of hosts. Such hosts include, but are not limited to, human, murine, rabbit, goat, guinea pig, camel, horse, mouse, rat, hamster, pig, micro-pig, chicken, goat, cow, sheep, dog, cat, non-human primate, and human. In specific embodiments, the host is a mouse, rabbit, goat, guinea pig, chicken, rat, hamster, pig, sheep, dog or cat. In preferred embodiments, the host is a mammal. In most preferred embodiments, the host is a human.

Other Preferred Embodiments

Other preferred embodiments of the claimed invention include an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least about 50 contiguous nucleotides in the nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in ATCC™ Deposit No:Z.

Also preferred is a nucleic acid molecule wherein said sequence of contiguous nucleotides is included in the nucleotide sequence of the portion of SEQ ID NO:X as defined in column 5, “ORF (From-To)”, in Table 1B.

Also preferred is a nucleic acid molecule wherein said sequence of contiguous nucleotides is included in the nucleotide sequence of the portion of SEQ ID NO:X as defined in columns 8 and 9, “NT From” and “NT To” respectively, in Table 2.

Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least about 150 contiguous nucleotides in the nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in ATCC™ Deposit No:Z.

Further preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least about 500 contiguous nucleotides in the nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in ATCC™ Deposit No:Z.

A further preferred embodiment is a nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the nucleotide sequence of the portion of SEQ ID NO:X defined in column 5, “ORF (From-To)”, in Table 1B.

A further preferred embodiment is a nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the nucleotide sequence of the portion of SEQ ID NO:X defined in columns 8 and 9, “NT From” and “NT To”, respectively, in Table 2.

A further preferred embodiment is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the complete nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in ATCC™ Deposit No:Z.

Also preferred is an isolated nucleic acid molecule which hybridizes under stringent hybridization conditions to a nucleic acid molecule comprising a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in ATCC™ Deposit No:Z, wherein said nucleic acid molecule which hybridizes does not hybridize under stringent hybridization conditions to a nucleic acid molecule having a nucleotide sequence consisting of only A residues or of only T residues.

Also preferred is a composition of matter comprising a DNA molecule which comprises the cDNA contained in ATCC™ Deposit No:Z.

Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least 50 contiguous nucleotides of the cDNA sequence contained in ATCC™ Deposit No:Z.

Also preferred is an isolated nucleic acid molecule, wherein said sequence of at least 50 contiguous nucleotides is included in the nucleotide sequence of an open reading frame sequence encoded by cDNA contained in ATCC™ Deposit No:Z.

Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to sequence of at least 150 contiguous nucleotides in the nucleotide sequence encoded by cDNA contained in ATCC™ Deposit No:Z.

A further preferred embodiment is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to sequence of at least 500 contiguous nucleotides in the nucleotide sequence encoded by cDNA contained in ATCC™ Deposit No:Z.

A further preferred embodiment is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the complete nucleotide sequence encoded by cDNA contained in ATCC™ Deposit No:Z.

A further preferred embodiment is a method for detecting in a biological sample a nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence encoded by cDNA contained in ATCC™ Deposit No:Z; which method comprises a step of comparing a nucleotide sequence of at least one nucleic acid molecule in said sample with a sequence selected from said group and determining whether the sequence of said nucleic acid molecule in said sample is at least 95% identical to said selected sequence.

Also preferred is the above method wherein said step of comparing sequences comprises determining the extent of nucleic acid hybridization between nucleic acid molecules in said sample and a nucleic acid molecule comprising said sequence selected from said group. Similarly, also preferred is the above method wherein said step of comparing sequences is performed by comparing the nucleotide sequence determined from a nucleic acid molecule in said sample with said sequence selected from said group. The nucleic acid molecules can comprise DNA molecules or RNA molecules.

A further preferred embodiment is a method for identifying the species, tissue or cell type of a biological sample which method comprises a step of detecting nucleic acid molecules in said sample, if any, comprising a nucleotide sequence that is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence of the cDNA contained in ATCC™ Deposit No:Z.

The method for identifying the species, tissue or cell type of a biological sample can comprise a step of detecting nucleic acid molecules comprising a nucleotide sequence in a panel of at least two nucleotide sequences, wherein at least one sequence in said panel is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from said group.

Also preferred is a method for diagnosing in a subject a pathological condition associated with abnormal structure or expression of a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto; or the cDNA contained in ATCC™ Deposit No:Z which encodes a protein, wherein the method comprises a step of detecting in a biological sample obtained from said subject nucleic acid molecules, if any, comprising a nucleotide sequence that is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence of cDNA contained in ATCC™ Deposit No:Z.

The method for diagnosing a pathological condition can comprise a step of detecting nucleic acid molecules comprising a nucleotide sequence in a panel of at least two nucleotide sequences, wherein at least one sequence in said panel is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from said group.

Also preferred is a composition of matter comprising isolated nucleic acid molecules wherein the nucleotide sequences of said nucleic acid molecules comprise a panel of at least two nucleotide sequences, wherein at least one sequence in said panel is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in Table 1B or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence encoded by cDNA contained in ATCC™ Deposit No:Z. The nucleic acid molecules can comprise DNA molecules or RNA molecules.

Also preferred is a composition of matter comprising isolated nucleic acid molecules wherein the nucleotide sequences of said nucleic acid molecules comprise a DNA microarray or “chip” of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50, 100, 150, 200, 250, 300, 500, 1000, 2000, 3000, or 4000 nucleotide sequences, wherein at least one sequence in said DNA microarray or “chip” is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X wherein X is any integer as defined in Table 1A and/or 1B; and a nucleotide sequence encoded by a human cDNA clone identified by a cDNA “Clone ID” in Table 1A and/or 1B.

Also preferred is an isolated polypeptide comprising an amino acid sequence at least 90% identical to a sequence of at least about 10 contiguous amino acids in the polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z.

Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 30 contiguous amino acids in the amino acid sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z.

Further preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 100 contiguous amino acids in the amino acid sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z.

Further preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to the complete amino acid sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z.

Further preferred is an isolated polypeptide comprising an amino acid sequence at least 90% identical to a sequence of at least about 10 contiguous amino acids in the complete amino acid sequence of a polypeptide encoded by contained in ATCC™ Deposit No:Z.

Also preferred is a polypeptide wherein said sequence of contiguous amino acids is included in the amino acid sequence of a portion of said polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or the polypeptide sequence of SEQ ID NO:Y.

Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 30 contiguous amino acids in the amino acid sequence of a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 100 contiguous amino acids in the amino acid sequence of a polypeptide encoded by cDNA contained in ATCC™ Deposit No:Z.

Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to the amino acid sequence of a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

Further preferred is an isolated antibody which binds specifically to a polypeptide comprising an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: a polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

Further preferred is a method for detecting in a biological sample a polypeptide comprising an amino acid sequence which is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: a polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z; which method comprises a step of comparing an amino acid sequence of at least one polypeptide molecule in said sample with a sequence selected from said group and determining whether the sequence of said polypeptide molecule in said sample is at least 90% identical to said sequence of at least 10 contiguous amino acids.

Also preferred is the above method wherein said step of comparing an amino acid sequence of at least one polypeptide molecule in said sample with a sequence selected from said group comprises determining the extent of specific binding of polypeptides in said sample to an antibody which binds specifically to a polypeptide comprising an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: a polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

Also preferred is the above method wherein said step of comparing sequences is performed by comparing the amino acid sequence determined from a polypeptide molecule in said sample with said sequence selected from said group.

Also preferred is a method for identifying the species, tissue or cell type of a biological sample which method comprises a step of detecting polypeptide molecules in said sample, if any, comprising an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

Also preferred is the above method for identifying the species, tissue or cell type of a biological sample, which method comprises a step of detecting polypeptide molecules comprising an amino acid sequence in a panel of at least two amino acid sequences, wherein at least one sequence in said panel is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the above group.

Also preferred is a method for diagnosing in a subject a pathological condition associated with abnormal structure or expression of a nucleic acid sequence identified in Table 1A, 1B or Table 2 encoding a polypeptide, which method comprises a step of detecting in a biological sample obtained from said subject polypeptide molecules comprising an amino acid sequence in a panel of at least two amino acid sequences, wherein at least one sequence in said panel is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

In any of these methods, the step of detecting said polypeptide molecules includes using an antibody.

Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a nucleotide sequence encoding a polypeptide wherein said polypeptide comprises an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

Also preferred is an isolated nucleic acid molecule, wherein said nucleotide sequence encoding a polypeptide has been optimized for expression of said polypeptide in a prokaryotic host.

Also preferred is a polypeptide molecule, wherein said polypeptide comprises an amino acid sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z.

Further preferred is a method of making a recombinant vector comprising inserting any of the above isolated nucleic acid molecule into a vector. Also preferred is the recombinant vector produced by this method. Also preferred is a method of making a recombinant host cell comprising introducing the vector into a host cell, as well as the recombinant host cell produced by this method.

Also preferred is a method of making an isolated polypeptide comprising culturing this recombinant host cell under conditions such that said polypeptide is expressed and recovering said polypeptide. Also preferred is this method of making an isolated polypeptide, wherein said recombinant host cell is a eukaryotic cell and said polypeptide is a human protein comprising an amino acid sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in ATCC™ Deposit No:Z. The isolated polypeptide produced by this method is also preferred.

Also preferred is a method of treatment of an individual in need of an increased level of a protein activity, which method comprises administering to such an individual a Therapeutic comprising an amount of an isolated polypeptide, polynucleotide, immunogenic fragment or analogue thereof, binding agent, antibody, or antigen binding fragment of the claimed invention effective to increase the level of said protein activity in said individual.

Also preferred is a method of treatment of an individual in need of a decreased level of a protein activity, which method comprised administering to such an individual a Therapeutic comprising an amount of an isolated polypeptide, polynucleotide, immunogenic fragment or analogue thereof, binding agent, antibody, or antigen binding fragment of the claimed invention effective to decrease the level of said protein activity in said individual.

Also preferred is a method of treatment of an individual in need of a specific delivery of toxic compositions to diseased cells (e.g., tumors, leukemias or lymphomas), which method comprises administering to such an individual a Therapeutic comprising an amount of an isolated polypeptide of the invention, including, but not limited to a binding agent, or antibody of the claimed invention that are associated with toxin or cytotoxic prodrugs.

Having generally described the invention, the same will be more readily understood by reference to the following examples, which are provided by way of illustration and are not intended as limiting.

Description of Table 6

Table 6 summarizes some of the ATCC™ Deposits, Deposit dates, and ATCC™ designation numbers of deposits made with the ATCC™ in connection with the present application. These deposits were made in addition to those described in the Table 1A.

TABLE 6 ATCC ™ ATCC ™ Deposits Deposit Date Designation Number LP01, LP02, LP03, LP04, May 20, 1997 209059, 209060, 209061, LP05, LP06, LP07, LP08, 209062, 209063, 209064, LP09, LP10, LP11, 209065, 209066, 209067, 209068, 209069 LP12 Jan. 12, 1998 209579 LP13 Jan. 12, 1998 209578 LP14 Jul. 16, 1998 203067 LP15 Jul. 16, 1998 203068 LP16 Feb. 1, 1999 203609 LP17 Feb. 1, 1999 203610 LP20 Nov. 17, 1998 203485 LP21 Jun. 18, 1999 PTA-252 LP22 Jun. 18, 1999 PTA-253 LP23 Dec. 22, 1999 PTA-1081

EXAMPLES Example 1 Isolation of a Selected cDNA Clone from the Deposited Sample

Each ATCC™ Deposit No:Z is contained in a plasmid vector. Table 7 identifies the vectors used to construct the cDNA library from which each clone was isolated. In many cases, the vector used to construct the library is a phage vector from which a plasmid has been excised. The following correlates the related plasmid for each phage vector used in constructing the cDNA library. For example, where a particular clone is identified in Table 7 as being isolated in the vector “Lambda Zap,” the corresponding deposited clone is in “pBLUESCRIPT™.”

Vector Used to Construct Library Corresponding Deposited Plasmid Lambda Zap pBLUESCRIPT ™ (pBS) Uni-Zap XR pBLUESCRIPT ™ (pBS) Zap Express pBK lafmid BA plafmid BA pSport1 pSport1 pCMVSport 2.0 pCMVSport 2.0 pCMVSport 3.0 pCMVSport 3.0 pCR ® 2.1 pCR ® 2.1

Vectors Lambda Zap (U.S. Pat. Nos. 5,128,256 and 5,286,636), Uni-Zap XR (U.S. Pat. Nos. 5,128,256 and 5,286,636), Zap Express (U.S. Pat. Nos. 5,128,256 and 5,286,636), pBLUESCRIPT™ (pBS) (Short, J. M. et al., Nucleic Acids Res. 16:7583-7600 (1988); Alting-Mees, M. A. and Short, J. M., Nucleic Acids Res. 17:9494 (1989)) and pBK (Alting-Mees, M. A. et al., Strategies 5:58-61 (1992)) are commercially available from STRATAGENE™ Cloning Systems, Inc., 11011 N. Torrey Pines Road, La Jolla, Calif., 92037. pBS contains an ampicillin resistance gene and pBK contains a neomycin resistance gene. Both can be transformed into E. coli strain XL-1 Blue, also available from STRATAGENE™. pBS comes in 4 forms SK+, SK−, KS+ and KS. The S and K refers to the orientation of the polylinker to the T7 and T3 primer sequences which flank the polylinker region (“S” is for Sad and “K” is for KpnI which are the first sites on each respective end of the linker). “+” or “−” refer to the orientation of the f1 origin of replication (“on”), such that in one orientation, single stranded rescue initiated from the f1 on generates sense strand DNA and in the other, antisense.

Vectors pSport1, pCMVSport 2.0 and pCMVSport 3.0, were obtained from Life Technologies, Inc., P. O. Box 6009, Gaithersburg, Md. 20897. All Sport vectors contain an ampicillin resistance gene and may be transformed into E. coli strain DH10B, also available from Life Technologies. (See, for instance, Gruber, C. E., et al., Focus 15:59 (1993)). Vector lafmid BA (Bento Soares, Columbia University, NY) contains an ampicillin resistance gene and can be transformed into E. coli strain XL-1 Blue. Vector pCR®2.1, which is available from Invitrogen, 1600 Faraday Avenue, Carlsbad, Calif. 92008, contains an ampicillin resistance gene and may be transformed into E. coli strain DH10B, available from Life Technologies. (See, for instance, Clark, J. M., Nuc. Acids Res. 16:9677-9686 (1988) and Mead, D. et al., Bio/Technology 9: (1991)). Preferably, a polynucleotide of the present invention does not comprise the phage vector sequences identified for the particular clone in Table 7, as well as the corresponding plasmid vector sequences designated above.

The deposited material in the sample assigned the ATCC™ Deposit Number cited by reference to Table 1A, Table 2, Table 6 and Table 7 for any given cDNA clone also may contain one or more additional plasmids, each comprising a cDNA clone different from that given clone. Thus, deposits sharing the same ATCC™ Deposit Number contain at least a plasmid for each ATCC™ Deposit No:Z.

TABLE 7 ATCC ™ Libraries owned by Catalog Catalog Description Vector Deposit HUKA HUKB HUKC HUKD Human Uterine Cancer Lambda ZAP II LP01 HUKE HUKF HUKG HCNA HCNB Human Colon Lambda Zap II LP01 HFFA Human Fetal Brain, random primed Lambda Zap II LP01 HTWA Resting T-Cell Lambda ZAP II LP01 HBQA Early Stage Human Brain, random Lambda ZAP II LP01 primed HLMB HLMF HLMG HLMH breast lymph node CDNA library Lambda ZAP II LP01 HLMI HLMJ HLMM HLMN HCQA HCQB human colon cancer Lamda ZAP II LP01 HMEA HMEC HMED HMEE Human Microvascular Endothelial Lambda ZAP II LP01 HMEF HMEG HMEI HMEJ Cells, fract. A HMEK HMEL HUSA HUSC Human Umbilical Vein Endothelial Lambda ZAP II LP01 Cells, fract. A HLQA HLQB Hepatocellular Tumor Lambda ZAP II LP01 HHGA HHGB HHGC HHGD Hemangiopericytoma Lambda ZAP II LP01 HSDM Human Striatum Depression, re- Lambda ZAP II LP01 rescue HUSH H Umbilical Vein Endothelial Cells, Lambda ZAP II LP01 frac A, re-excision HSGS Salivary gland, subtracted Lambda ZAP II LP01 HFXA HFXB HFXC HFXD HFXE Brain frontal cortex Lambda ZAP II LP01 HFXF HFXG HFXH HPQA HPQB HPQC PERM TF274 Lambda ZAP II LP01 HFXJ HFXK Brain Frontal Cortex, re-excision Lambda ZAP II LP01 HCWA HCWB HCWC HCWD CD34 positive cells (Cord Blood) ZAP Express LP02 HCWE HCWF HCWG HCWH HCWI HCWJ HCWK HCUA HCUB HCUC CD34 depleted Buffy Coat (Cord ZAP Express LP02 Blood) HRSM A-14 cell line ZAP Express LP02 HRSA A1-CELL LINE ZAP Express LP02 HCUD HCUE HCUF HCUG CD34 depleted Buffy Coat (Cord ZAP Express LP02 HCUH HCUI Blood), re-excision HBXE HBXF HBXG H. Whole Brain #2, re-excision ZAP Express LP02 HRLM L8 cell line ZAP Express LP02 HBXA HBXB HBXC HBXD Human Whole Brain #2 - Oligo dT ZAP Express LP02 >1.5 Kb HUDA HUDB HUDC Testes ZAP Express LP02 HHTM HHTN HHTO H. hypothalamus, frac A; re-excision ZAP Express LP02 HHTL H. hypothalamus, frac A ZAP Express LP02 HASA HASD Human Adult Spleen Uni-ZAP XR LP03 HFKC HFKD HFKE HFKF HFKG Human Fetal Kidney Uni-ZAP XR LP03 HE8A HE8B HE8C HE8D HE8E Human 8 Week Whole Embryo Uni-ZAP XR LP03 HE8F HE8M HE8N HGBA HGBD HGBE HGBF Human Gall Bladder Uni-ZAP XR LP03 HGBG HGBH HGBI HLHA HLHB HLHC HLHD HLHE Human Fetal Lung III Uni-ZAP XR LP03 HLHF HLHG HLHH HLHQ HPMA HPMB HPMC HPMD Human Placenta Uni-ZAP XR LP03 HPME HPMF HPMG HPMH HPRA HPRB HPRC HPRD Human Prostate Uni-ZAP XR LP03 HSIA HSIC HSID HSIE Human Adult Small Intestine Uni-ZAP XR LP03 HTEA HTEB HTEC HTED HTEE Human Testes Uni-ZAP XR LP03 HTEF HTEG HTEH HTEI HTEJ HTEK HTPA HTPB HTPC HTPD HTPE Human Pancreas Tumor Uni-ZAP XR LP03 HTTA HTTB HTTC HTTD HTTE Human Testes Tumor Uni-ZAP XR LP03 HTTF HAPA HAPB HAPC HAPM Human Adult Pulmonary Uni-ZAP XR LP03 HETA HETB HETC HETD HETE Human Endometrial Tumor Uni-ZAP XR LP03 HETF HETG HETH HETI HHFB HHFC HHFD HHFE HHFF Human Fetal Heart Uni-ZAP XR LP03 HHFG HHFH HHFI HHPB HHPC HHPD HHPE HHPF Human Hippocampus Uni-ZAP XR LP03 HHPG HHPH HCE1 HCE2 HCE3 HCE4 HCE5 Human Cerebellum Uni-ZAP XR LP03 HCEB HCEC HCED HCEE HCEF HCEG HUVB HUVC HUVD HUVE Human Umbilical Vein, Endo. Uni-ZAP XR LP03 remake HSTA HSTB HSTC HSTD Human Skin Tumor Uni-ZAP XR LP03 HTAA HTAB HTAC HTAD HTAE Human Activated T-Cells Uni-ZAP XR LP03 HFEA HFEB HFEC Human Fetal Epithelium (Skin) Uni-ZAP XR LP03 HJPA HJPB HJPC HJPD HUMAN JURKAT MEMBRANE Uni-ZAP XR LP03 BOUND POLYSOMES HESA Human epithelioid sarcoma Uni-Zap XR LP03 HLTA HLTB HLTC HLTD HLTE Human T-Cell Lymphoma Uni-ZAP XR LP03 HLTF HFTA HFTB HFTC HFTD Human Fetal Dura Mater Uni-ZAP XR LP03 HRDA HRDB HRDC HRDD Human Rhabdomyosarcoma Uni-ZAP XR LP03 HRDE HRDF HCAA HCAB HCAC Cem cells cyclohexamide treated Uni-ZAP XR LP03 HRGA HRGB HRGC HRGD Raji Cells, cyclohexamide treated Uni-ZAP XR LP03 HSUA HSUB HSUC HSUM Supt Cells, cyclohexamide treated Uni-ZAP XR LP03 HT4A HT4C HT4D Activated T-Cells, 12 hrs. Uni-ZAP XR LP03 HE9A HE9B HE9C HE9D HE9E Nine Week Old Early Stage Human Uni-ZAP XR LP03 HE9F HE9G HE9H HE9M HE9N HATA HATB HATC HATD HATE Human Adrenal Gland Tumor Uni-ZAP XR LP03 HT5A Activated T-Cells, 24 hrs. Uni-ZAP XR LP03 HFGA HFGM Human Fetal Brain Uni-ZAP XR LP03 HNEA HNEB HNEC HNED HNEE Human Neutrophil Uni-ZAP XR LP03 HBGB HBGD Human Primary Breast Cancer Uni-ZAP XR LP03 HBNA HBNB Human Normal Breast Uni-ZAP XR LP03 HCAS Cem Cells, cyclohexamide treated, Uni-ZAP XR LP03 subtra HHPS Human Hippocampus, subtracted pBS LP03 HKCS HKCU Human Colon Cancer, subtracted pBS LP03 HRGS Raji cells, cyclohexamide treated, pBS LP03 subtracted HSUT Supt cells, cyclohexamide treated, pBS LP03 differentially expressed HT4S Activated T-Cells, 12 hrs, Uni-ZAP XR LP03 subtracted HCDA HCDB HCDC HCDD Human Chondrosarcoma Uni-ZAP XR LP03 HCDE HOAA HOAB HOAC Human Osteosarcoma Uni-ZAP XR LP03 HTLA HTLB HTLC HTLD HTLE Human adult testis, large inserts Uni-ZAP XR LP03 HTLF HLMA HLMC HLMD Breast Lymph node cDNA library Uni-ZAP XR LP03 H6EA H6EB H6EC HL-60, PMA 4H Uni-ZAP XR LP03 HTXA HTXB HTXC HTXD HTXE Activated T-Cell (12 hs)/Thiouridine Uni-ZAP XR LP03 HTXF HTXG HTXH labelledEco HNFA HNFB HNFC HNFD HNFE Human Neutrophil, Activated Uni-ZAP XR LP03 HNFF HNFG HNFH HNFJ HTOB HTOC HUMAN TONSILS, FRACTION 2 Uni-ZAP XR LP03 HMGB Human OB MG63 control fraction I Uni-ZAP XR LP03 HOPB Human OB HOS control fraction I Uni-ZAP XR LP03 HORB Human OB HOS treated (10 nM Uni-ZAP XR LP03 E2) fraction I HSVA HSVB HSVC Human Chronic Synovitis Uni-ZAP XR LP03 HROA HUMAN STOMACH Uni-ZAP XR LP03 HBJA HBJB HBJC HBJD HBJE HUMAN B CELL LYMPHOMA Uni-ZAP XR LP03 HBJF HBJG HBJH HBJI HBJJ HBJK HCRA HCRB HCRC human corpus colosum Uni-ZAP XR LP03 HODA HODB HODC HODD human ovarian cancer Uni-ZAP XR LP03 HDSA Dermatofibrosarcoma Protuberance Uni-ZAP XR LP03 HMWA HMWB HMWC HMWD Bone Marrow Cell Line (RS4; 11) Uni-ZAP XR LP03 HMWE HMWF HMWG HMWH HMWI HMWJ HSOA stomach cancer (human) Uni-ZAP XR LP03 HERA SKIN Uni-ZAP XR LP03 HMDA Brain-medulloblastoma Uni-ZAP XR LP03 HGLA HGLB HGLD Glioblastoma Uni-ZAP XR LP03 HEAA H. Atrophic Endometrium Uni-ZAP XR LP03 HBCA HBCB H. Lymph node breast Cancer Uni-ZAP XR LP03 HPWT Human Prostate BPH, re-excision Uni-ZAP XR LP03 HFVG HFVH HFVI Fetal Liver, subtraction II pBS LP03 HNFI Human Neutrophils, Activated, re- pBS LP03 excision HBMB HBMC HBMD Human Bone Marrow, re-excision pBS LP03 HKML HKMM HKMN H. Kidney Medulla, re-excision pBS LP03 HKIX HKIY H. Kidney Cortex, subtracted pBS LP03 HADT H. Amygdala Depression, pBS LP03 subtracted H6AS Hl-60, untreated, subtracted Uni-ZAP XR LP03 H6ES HL-60, PMA 4H, subtracted Uni-ZAP XR LP03 H6BS HL-60, RA 4 h, Subtracted Uni-ZAP XR LP03 H6CS HL-60, PMA 1 d, subtracted Uni-ZAP XR LP03 HTXJ HTXK Activated T-cell(12 h)/Thiouridine- Uni-ZAP XR LP03 re-excision HMSA HMSB HMSC HMSD Monocyte activated Uni-ZAP XR LP03 HMSE HMSF HMSG HMSH HMSI HMSJ HMSK HAGA HAGB HAGC HAGD Human Amygdala Uni-ZAP XR LP03 HAGE HAGF HSRA HSRB HSRE STROMAL-OSTEOCLASTOMA Uni-ZAP XR LP03 HSRD HSRF HSRG HSRH Human Osteoclastoma Stromal Uni-ZAP XR LP03 Cells - unamplified HSQA HSQB HSQC HSQD HSQE Stromal cell TF274 Uni-ZAP XR LP03 HSQF HSQG HSKA HSKB HSKC HSKD HSKE Smooth muscle, serum treated Uni-ZAP XR LP03 HSKF HSKZ HSLA HSLB HSLC HSLD HSLE Smooth muscle, control Uni-ZAP XR LP03 HSLF HSLG HSDA HSDD HSDE HSDF HSDG Spinal cord Uni-ZAP XR LP03 HSDH HPWS Prostate-BPH subtracted II pBS LP03 HSKW HSKX HSKY Smooth Muscle-HASTE pBS LP03 normalized HFPB HFPC HFPD H. Frontal cortex, epileptic; re- Uni-ZAP XR LP03 excision HSDI HSDJ HSDK Spinal Cord, re-excision Uni-ZAP XR LP03 HSKN HSKO Smooth Muscle Serum Treated, pBS LP03 Norm HSKG HSKH HSKI Smooth muscle, serum induced, re- pBS LP03 exc HFCA HFCB HFCC HFCD HFCE Human Fetal Brain Uni-ZAP XR LP04 HFCF HPTA HPTB HPTD Human Pituitary Uni-ZAP XR LP04 HTHB HTHC HTHD Human Thymus Uni-ZAP XR LP04 HE6B HE6C HE6D HE6E HE6F Human Whole Six Week Old Uni-ZAP XR LP04 HE6G HE6S Embryo HSSA HSSB HSSC HSSD HSSE Human Synovial Sarcoma Uni-ZAP XR LP04 HSSF HSSG HSSH HSSI HSSJ HSSK HE7T 7 Week Old Early Stage Human, Uni-ZAP XR LP04 subtracted HEPA HEPB HEPC Human Epididymus Uni-ZAP XR LP04 HSNA HSNB HSNC HSNM HSNN Human Synovium Uni-ZAP XR LP04 HPFB HPFC HPFD HPFE Human Prostate Cancer, Stage C Uni-ZAP XR LP04 fraction HE2A HE2D HE2E HE2H HE2I 12 Week Old Early Stage Human Uni-ZAP XR LP04 HE2M HE2N HE2O HE2B HE2C HE2F HE2G HE2P 12 Week Old Early Stage Human, II Uni-ZAP XR LP04 HE2Q HPTS HPTT HPTU Human Pituitary, subtracted Uni-ZAP XR LP04 HAUA HAUB HAUC Amniotic Cells - TNF induced Uni-ZAP XR LP04 HAQA HAQB HAQC HAQD Amniotic Cells - Primary Culture Uni-ZAP XR LP04 HWTA HWTB HWTC wilm's tumor Uni-ZAP XR LP04 HBSD Bone Cancer, re-excision Uni-ZAP XR LP04 HSGB Salivary gland, re-excision Uni-ZAP XR LP04 HSJA HSJB HSJC Smooth muscle-ILb induced Uni-ZAP XR LP04 HSXA HSXB HSXC HSXD Human Substantia Nigra Uni-ZAP XR LP04 HSHA HSHB HSHC Smooth muscle, IL1b induced Uni-ZAP XR LP04 HOUA HOUB HOUC HOUD Adipocytes Uni-ZAP XR LP04 HOUE HPWA HPWB HPWC HPWD Prostate BPH Uni-ZAP XR LP04 HPWE HELA HELB HELC HELD HELE Endothelial cells-control Uni-ZAP XR LP04 HELF HELG HELH HEMA HEMB HEMC HEMD Endothelial-induced Uni-ZAP XR LP04 HEME HEMF HEMG HEMH HBIA HBIB HBIC Human Brain, Striatum Uni-ZAP XR LP04 HHSA HHSB HHSC HHSD HHSE Human Hypothalmus, Schizophrenia Uni-ZAP XR LP04 HNGA HNGB HNGC HNGD neutrophils control Uni-ZAP XR LP04 HNGE HNGF HNGG HNGH HNGI HNGJ HNHA HNHB HNHC HNHD Neutrophils IL-1 and LPS induced Uni-ZAP XR LP04 HNHE HNHF HNHG HNHH HNHI HNHJ HSDB HSDC STRIATUM DEPRESSION Uni-ZAP XR LP04 HHPT Hypothalamus Uni-ZAP XR LP04 HSAT HSAU HSAV HSAW HSAX Anergic T-cell Uni-ZAP XR LP04 HSAY HSAZ HBMS HBMT HBMU HBMV Bone marrow Uni-ZAP XR LP04 HBMW HBMX HOEA HOEB HOEC HOED HOEE Osteoblasts Uni-ZAP XR LP04 HOEF HOEJ HAIA HAIB HAIC HAID HAIE Epithelial-TNFa and INF induced Uni-ZAP XR LP04 HAIF HTGA HTGB HTGC HTGD Apoptotic T-cell Uni-ZAP XR LP04 HMCA HMCB HMCC HMCD Macrophage-oxLDL Uni-ZAP XR LP04 HMCE HMAA HMAB HMAC HMAD Macrophage (GM-CSF treated) Uni-ZAP XR LP04 HMAE HMAF HMAG HPHA Normal Prostate Uni-ZAP XR LP04 HPIA HPIB HPIC LNCAP prostate cell line Uni-ZAP XR LP04 HPJA HPJB HPJC PC3 Prostate cell line Uni-ZAP XR LP04 HOSE HOSF HOSG Human Osteoclastoma, re-excision Uni-ZAP XR LP04 HTGE HTGF Apoptotic T-cell, re-excision Uni-ZAP XR LP04 HMAJ HMAK H Macrophage (GM-CSF treated), Uni-ZAP XR LP04 re-excision HACB HACC HACD Human Adipose Tissue, re-excision Uni-ZAP XR LP04 HFPA H. Frontal Cortex, Epileptic Uni-ZAP XR LP04 HFAA HFAB HFAC HFAD HFAE Alzheimer's, spongy change Uni-ZAP XR LP04 HFAM Frontal Lobe, Dementia Uni-ZAP XR LP04 HMIA HMIB HMIC Human Manic Depression Tissue Uni-ZAP XR LP04 HTSA HTSE HTSF HTSG HTSH Human Thymus pBS LP05 HPBA HPBB HPBC HPBD HPBE Human Pineal Gland pBS LP05 HSAA HSAB HSAC HSA 172 Cells pBS LP05 HSBA HSBB HSBC HSBM HSC172 cells pBS LP05 HJAA HJAB HJAC HJAD Jurkat T-cell G1 phase pBS LP05 HJBA HJBB HJBC HJBD Jurkat T-Cell, S phase pBS LP05 HAFA HAFB Aorta endothelial cells + TNF-a pBS LP05 HAWA HAWB HAWC Human White Adipose pBS LP05 HTNA HTNB Human Thyroid pBS LP05 HONA Normal Ovary, Premenopausal pBS LP05 HARA HARB Human Adult Retina pBS LP05 HLJA HLJB Human Lung pCMVSport 1 LP06 HOFM HOFN HOFO H. Ovarian Tumor, II, OV5232 pCMVSport 2.0 LP07 HOGA HOGB HOGC OV 10-3-95 pCMVSport 2.0 LP07 HCGL CD34+cells, II pCMVSport 2.0 LP07 HDLA Hodgkin's Lymphoma I pCMVSport 2.0 LP07 HDTA HDTB HDTC HDTD HDTE Hodgkin's Lymphoma II pCMVSport 2.0 LP07 HKAA HKAB HKAC HKAD Keratinocyte pCMVSport 2.0 LP07 HKAE HKAF HKAG HKAH HCIM CAPFINDER, Crohn's Disease, lib 2 pCMVSport 2.0 LP07 HKAL Keratinocyte, lib 2 pCMVSport 2.0 LP07 HKAT Keratinocyte, lib 3 pCMVSport 2.0 LP07 HNDA Nasal polyps pCMVSport 2.0 LP07 HDRA H. Primary Dendritic Cells, lib 3 pCMVSport 2.0 LP07 HOHA HOHB HOHC Human Osteoblasts II pCMVSport 2.0 LP07 HLDA HLDB HLDC Liver, Hepatoma pCMVSport 3.0 LP08 HLDN HLDO HLDP Human Liver, normal pCMVSport 3.0 LP08 HMTA pBMC stimulated w/ poly I/C pCMVSport 3.0 LP08 HNTA NTERA2, control pCMVSport 3.0 LP08 HDPA HDPB HDPC HDPD HDPF Primary Dendritic Cells, lib 1 pCMVSport 3.0 LP08 HDPG HDPH HDPI HDPJ HDPK HDPM HDPN HDPO HDPP Primary Dendritic cells, frac 2 pCMVSport 3.0 LP08 HMUA HMUB HMUC Myoloid Progenitor Cell Line pCMVSport 3.0 LP08 HHEA HHEB HHEC HHED T Cell helper I pCMVSport 3.0 LP08 HHEM HHEN HHEO HHEP T cell helper II pCMVSport 3.0 LP08 HEQA HEQB HEQC Human endometrial stromal cells pCMVSport 3.0 LP08 HJMA HJMB Human endometrial stromal cells- pCMVSport 3.0 LP08 treated with progesterone HSWA HSWB HSWC Human endometrial stromal cells- pCMVSport 3.0 LP08 treated with estradiol HSYA HSYB HSYC Human Thymus Stromal Cells pCMVSport 3.0 LP08 HLWA HLWB HLWC Human Placenta pCMVSport 3.0 LP08 HRAA HRAB HRAC Rejected Kidney, lib 4 pCMVSport 3.0 LP08 HMTM PCR, pBMC I/C treated PCRII LP09 HMJA H. Meniingima, M6 pSport 1 LP10 HMKA HMKB HMKC HMKD H. Meningima, M1 pSport 1 LP10 HMKE HUSG HUSI Human umbilical vein endothelial pSport 1 LP10 cells, IL-4 induced HUSX HUSY Human Umbilical Vein Endothelial pSport 1 LP10 Cells, uninduced HOFA Ovarian Tumor I, OV5232 pSport 1 LP10 HCFA HCFB HCFC HCFD T-Cell PHA 16 hrs pSport 1 LP10 HCFL HCFM HCFN HCFO T-Cell PHA 24 hrs pSport 1 LP10 HADA HADC HADD HADE Human Adipose pSport 1 LP10 HADF HADG HOVA HOVB HOVC Human Ovary pSport 1 LP10 HTWB HTWC HTWD HTWE Resting T-Cell Library, II pSport 1 LP10 HTWF HMMA Spleen metastic melanoma pSport 1 LP10 HLYA HLYB HLYC HLYD HLYE Spleen, Chronic lymphocytic pSport 1 LP10 leukemia HCGA CD34+ cell, I pSport 1 LP10 HEOM HEON Human Eosinophils pSport 1 LP10 HTDA Human Tonsil, Lib 3 pSport 1 LP10 HSPA Salivary Gland, Lib 2 pSport 1 LP10 HCHA HCHB HCHC Breast Cancer cell line, MDA 36 pSport 1 LP10 HCHM HCHN Breast Cancer Cell line, angiogenic pSport 1 LP10 HCIA Crohn's Disease pSport 1 LP10 HDAA HDAB HDAC HEL cell line pSport 1 LP10 HABA Human Astrocyte pSport 1 LP10 HUFA HUFB HUFC Ulcerative Colitis pSport 1 LP10 HNTM NTERA2 + retinoic acid, 14 days pSport 1 LP10 HDQA Primary Dendritic cells, CapFinder2, pSport 1 LP10 frac 1 HDQM Primary Dendritic Cells, CapFinder, pSport 1 LP10 frac 2 HLDX Human Liver, normal, CapFinder pSport 1 LP10 HULA HULB HULC Human Dermal Endothelial pSport 1 LP10 Cells, untreated HUMA Human Dermal Endothelial pSport 1 LP10 cells, treated HCJA Human Stromal Endometrial pSport 1 LP10 fibroblasts, untreated HCJM Human Stromal endometrial pSport 1 LP10 fibroblasts, treated w/ estradiol HEDA Human Stromal endometrial pSport 1 LP10 fibroblasts, treated with progesterone HFNA Human ovary tumor cell OV350721 pSport 1 LP10 HKGA HKGB HKGC HKGD Merkel Cells pSport 1 LP10 HISA HISB HISC Pancreas Islet Cell Tumor pSport 1 LP10 HLSA Skin, burned pSport 1 LP10 HBZA Prostate, BPH, Lib 2 pSport 1 LP10 HBZS Prostate BPH, Lib 2, subtracted pSport 1 LP10 HFIA HFIB HFIC Synovial Fibroblasts (control) pSport 1 LP10 HFIH HFII HFIJ Synovial hypoxia pSport 1 LP10 HFIT HFIU HFIV Synovial IL-1/TNF stimulated pSport 1 LP10 HGCA Messangial cell, frac 1 pSport 1 LP10 HMVA HMVB HMVC Bone Marrow Stromal Cell, pSport 1 LP10 untreated HFIX HFIY HFIZ Synovial Fibroblasts (Il1/TNF), subt pSport 1 LP10 HFOX HFOY HFOZ Synovial hypoxia-RSF subtracted pSport 1 LP10 HMQA HMQB HMQC HMQD Human Activated Monocytes Uni-ZAP XR LP11 HLIA HLIB HLIC Human Liver pCMVSport 1 LP012 HHBA HHBB HHBC HHBD Human Heart pCMVSport 1 LP012 HHBE HBBA HBBB Human Brain pCMVSport 1 LP012 HLJA HLJB HLJC HLJD HLJE Human Lung pCMVSport 1 LP012 HOGA HOGB HOGC Ovarian Tumor pCMVSport 2.0 LP012 HTJM Human Tonsils, Lib 2 pCMVSport 2.0 LP012 HAMF HAMG KMH2 pCMVSport 3.0 LP012 HAJA HAJB HAJC L428 pCMVSport 3.0 LP012 HWBA HWBB HWBC HWBD Dendritic cells, pooled pCMVSport 3.0 LP012 HWBE HWAA HWAB HWAC HWAD Human Bone Marrow, treated pCMVSport 3.0 LP012 HWAE HYAA HYAB HYAC B Cell lymphoma pCMVSport 3.0 LP012 HWHG HWHH HWHI Healing groin wound, 6.5 hours post pCMVSport 3.0 LP012 incision HWHP HWHQ HWHR Healing groin wound; 7.5 hours pCMVSport 3.0 LP012 post incision HARM Healing groin wound - zero hr post- pCMVSport 3.0 LP012 incision (control) HBIM Olfactory epithelium; nasalcavity pCMVSport 3.0 LP012 HWDA Healing Abdomen wound; 70&90 min pCMVSport 3.0 LP012 post incision HWEA Healing Abdomen Wound; 15 days pCMVSport 3.0 LP012 post incision HWJA Healing Abdomen Wound; 21&29 pCMVSport 3.0 LP012 days HNAL Human Tongue, frac 2 pSport 1 LP012 HMJA H. Meniingima, M6 pSport 1 LP012 HMKA HMKB HMKC HMKD H. Meningima, M1 pSport 1 LP012 HMKE HOFA Ovarian Tumor I, OV5232 pSport 1 LP012 HCFA HCFB HCFC HCFD T-Cell PHA 16 hrs pSport 1 LP012 HCFL HCFM HCFN HCFO T-Cell PHA 24 hrs pSport 1 LP012 HMMA HMMB HMMC Spleen metastic melanoma pSport 1 LP012 HTDA Human Tonsil, Lib 3 pSport 1 LP012 HDBA Human Fetal Thymus pSport 1 LP012 HDUA Pericardium pSport 1 LP012 HBZA Prostate, BPH, Lib 2 pSport 1 LP012 HWCA Larynx tumor pSport 1 LP012 HWKA Normal lung pSport 1 LP012 HSMB Bone marrow stroma, treated pSport 1 LP012 HBHM Normal trachea pSport 1 LP012 HLFC Human Larynx pSport 1 LP012 HLRB Siebben Polyposis pSport 1 LP012 HNIA Mammary Gland pSport 1 LP012 HNJB Palate carcinoma pSport 1 LP012 HNKA Palate normal pSport 1 LP012 HMZA Pharynx carcinoma pSport 1 LP012 HABG Cheek Carcinoma pSport 1 LP012 HMZM Pharynx Carcinoma pSport 1 LP012 HDRM Larynx Carcinoma pSport 1 LP012 HVAA Pancreas normal PCA4 No pSport 1 LP012 HICA Tongue carcinoma pSport 1 LP012 HUKA HUKB HUKC HUKD Human Uterine Cancer Lambda ZAP II LP013 HUKE HFFA Human Fetal Brain, random primed Lambda ZAP II LP013 HTUA Activated T-cell labeled with 4- Lambda ZAP II LP013 thioluri HBQA Early Stage Human Brain, random Lambda ZAP II LP013 primed HMEB Human microvascular Endothelial Lambda ZAP II LP013 cells, fract. B HUSH Human Umbilical Vein Endothelial Lambda ZAP II LP013 cells, fract. A, re-excision HLQC HLQD Hepatocellular tumor, re-excision Lambda ZAP II LP013 HTWJ HTWK HTWL Resting T-cell, re-excision Lambda ZAP II LP013 HF6S Human Whole 6 week Old Embryo pBLUESCRIPT ™ LP013 (II), subt HHPS Human Hippocampus, subtracted pBLUESCRIPT ™ LP013 HL1S LNCAP, differential expression pBLUESCRIPT ™ LP013 HLHS HLHT Early Stage Human Lung, pBLUESCRIPT ™ LP013 Subtracted HSUS Supt cells, cyclohexamide treated, pBLUESCRIPT ™ LP013 subtracted HSUT Supt cells, cyclohexamide treated, pBLUESCRIPT ™ LP013 differentially expressed HSDS H. Striatum Depression, subtracted pBLUESCRIPT ™ LP013 HPTZ Human Pituitary, Subtracted VII pBLUESCRIPT ™ LP013 HSDX H. Striatum Depression, subt II pBLUESCRIPT ™ LP013 HSDZ H. Striatum Depression, subt pBLUESCRIPT ™ LP013 HPBA HPBB HPBC HPBD HPBE Human Pineal Gland pBLUESCRIPT ™ SK− LP013 HRTA Colorectal Tumor pBLUESCRIPT ™ SK− LP013 HSBA HSBB HSBC HSBM HSC172 cells pBLUESCRIPT ™ SK− LP013 HJAA HJAB HJAC HJAD Jurkat T-cell G1 phase pBLUESCRIPT ™ SK− LP013 HJBA HJBB HJBC HJBD Jurkat T-cell, S1 phase pBLUESCRIPT ™ SK− LP013 HTNA HTNB Human Thyroid pBLUESCRIPT ™ SK− LP013 HAHA HAHB Human Adult Heart Uni-ZAP XR LP013 HE6A Whole 6 week Old Embryo Uni-ZAP XR LP013 HFCA HFCB HFCC HFCD HFCE Human Fetal Brain Uni-ZAP XR LP013 HFKC HFKD HFKE HFKF HFKG Human Fetal Kidney Uni-ZAP XR LP013 HGBA HGBD HGBE HGBF Human Gall Bladder Uni-ZAP XR LP013 HGBG HPRA HPRB HPRC HPRD Human Prostate Uni-ZAP XR LP013 HTEA HTEB HTEC HTED HTEE Human Testes Uni-ZAP XR LP013 HTTA HTTB HTTC HTTD HTTE Human Testes Tumor Uni-ZAP XR LP013 HYBA HYBB Human Fetal Bone Uni-ZAP XR LP013 HFLA Human Fetal Liver Uni-ZAP XR LP013 HHFB HHFC HHFD HHFE HHFF Human Fetal Heart Uni-ZAP XR LP013 HUVB HUVC HUVD HUVE Human Umbilical Vein, End. Uni-ZAP XR LP013 remake HTHB HTHC HTHD Human Thymus Uni-ZAP XR LP013 HSTA HSTB HSTC HSTD Human Skin Tumor Uni-ZAP XR LP013 HTAA HTAB HTAC HTAD HTAE Human Activated T-cells Uni-ZAP XR LP013 HFEA HFEB HFEC Human Fetal Epithelium (skin) Uni-ZAP XR LP013 HJPA HJPB HJPC HJPD Human Jurkat Membrane Bound Uni-ZAP XR LP013 Polysomes HESA Human Epithelioid Sarcoma Uni-ZAP XR LP013 HALS Human Adult Liver, Subtracted Uni-ZAP XR LP013 HFTA HFTB HFTC HFTD Human Fetal Dura Mater Uni-ZAP XR LP013 HCAA HCAB HCAC Cem cells, cyclohexamide treated Uni-ZAP XR LP013 HRGA HRGB HRGC HRGD Raji Cells, cyclohexamide treated Uni-ZAP XR LP013 HE9A HE9B HE9C HE9D HE9E Nine Week Old Early Stage Human Uni-ZAP XR LP013 HSFA Human Fibrosarcoma Uni-ZAP XR LP013 HATA HATB HATC HATD HATE Human Adrenal Gland Tumor Uni-ZAP XR LP013 HTRA Human Trachea Tumor Uni-ZAP XR LP013 HE2A HE2D HE2E HE2H HE2I 12 Week Old Early Stage Human Uni-ZAP XR LP013 HE2B HE2C HE2F HE2G HE2P 12 Week Old Early Stage Human, II Uni-ZAP XR LP013 HNEA HNEB HNEC HNED HNEE Human Neutrophil Uni-ZAP XR LP013 HBGA Human Primary Breast Cancer Uni-ZAP XR LP013 HPTS HPTT HPTU Human Pituitary, subtracted Uni-ZAP XR LP013 HMQA HMQB HMQC HMQD Human Activated Monocytes Uni-ZAP XR LP013 HOAA HOAB HOAC Human Osteosarcoma Uni-ZAP XR LP013 HTOA HTOD HTOE HTOF HTOG human tonsils Uni-ZAP XR LP013 HMGB Human OB MG63 control fraction I Uni-ZAP XR LP013 HOPB Human OB HOS control fraction I Uni-ZAP XR LP013 HOQB Human OB HOS treated (1 nM E2) Uni-ZAP XR LP013 fraction I HAUA HAUB HAUC Amniotic Cells - TNF induced Uni-ZAP XR LP013 HAQA HAQB HAQC HAQD Amniotic Cells - Primary Culture Uni-ZAP XR LP013 HROA HROC HUMAN STOMACH Uni-ZAP XR LP013 HBJA HBJB HBJC HBJD HBJE HUMAN B CELL LYMPHOMA Uni-ZAP XR LP013 HODA HODB HODC HODD human ovarian cancer Uni-ZAP XR LP013 HCPA Corpus Callosum Uni-ZAP XR LP013 HSOA stomach cancer (human) Uni-ZAP XR LP013 HERA SKIN Uni-ZAP XR LP013 HMDA Brain-medulloblastoma Uni-ZAP XR LP013 HGLA HGLB HGLD Glioblastoma Uni-ZAP XR LP013 HWTA HWTB HWTC wilm's tumor Uni-ZAP XR LP013 HEAA H. Atrophic Endometrium Uni-ZAP XR LP013 HAPN HAPO HAPP HAPQ HAPR Human Adult Pulmonary; re- Uni-ZAP XR LP013 excision HLTG HLTH Human T-cell lymphoma; re- Uni-ZAP XR LP013 excision HAHC HAHD HAHE Human Adult Heart; re-excision Uni-ZAP XR LP013 HAGA HAGB HAGC HAGD Human Amygdala Uni-ZAP XR LP013 HAGE HSJA HSJB HSJC Smooth muscle-ILb induced Uni-ZAP XR LP013 HSHA HSHB HSHC Smooth muscle, IL1b induced Uni-ZAP XR LP013 HPWA HPWB HPWC HPWD Prostate BPH Uni-ZAP XR LP013 HPWE HPIA HPIB HPIC LNCAP prostate cell line Uni-ZAP XR LP013 HPJA HPJB HPJC PC3 Prostate cell line Uni-ZAP XR LP013 HBTA Bone Marrow Stroma, TNF&LPS Uni-ZAP XR LP013 ind HMCF HMCG HMCH HMCI Macrophage-oxLDL; re-excision Uni-ZAP XR LP013 HMCJ HAGG HAGH HAGI Human Amygdala; re-excision Uni-ZAP XR LP013 HACA H. Adipose Tissue Uni-ZAP XR LP013 HKFB K562 + PMA (36 hrs), re-excision ZAP Express LP013 HCWT HCWU HCWV CD34 positive cells (cord blood), re- ZAP Express LP013 ex HBWA Whole brain ZAP Express LP013 HBXA HBXB HBXC HBXD Human Whole Brain #2 - Oligo dT ZAP Express LP013 >1.5 Kb HAVM Temporal cortex-Alzheizmer pT-Adv LP014 HAVT Hippocampus, Alzheimer pT-Adv LP014 Subtracted HHAS CHME Cell Line Uni-ZAP XR LP014 HAJR Larynx normal pSport 1 LP014 HWLE HWLF HWLG HWLH Colon Normal pSport 1 LP014 HCRM HCRN HCRO Colon Carcinoma pSport 1 LP014 HWLI HWLJ HWLK Colon Normal pSport 1 LP014 HWLQ HWLR HWLS HWLT Colon Tumor pSport 1 LP014 HBFM Gastrocnemius Muscle pSport 1 LP014 HBOD HBOE Quadriceps Muscle pSport 1 LP014 HBKD HBKE Soleus Muscle pSport 1 LP014 HCCM Pancreatic Langerhans pSport 1 LP014 HWGA Larynx carcinoma pSport 1 LP014 HWGM HWGN Larynx carcinoma pSport 1 LP014 HWLA HWLB HWLC Normal colon pSport 1 LP014 HWLM HWLN Colon Tumor pSport 1 LP014 HVAM HYAN HVAO Pancreas Tumor pSport 1 LP014 HWGQ Larynx carcinoma pSport 1 LP014 HAQM HAQN Salivary Gland pSport 1 LP014 HASM Stomach; normal pSport 1 LP014 HBCM Uterus; normal pSport 1 LP014 HCDM Testis; normal pSport 1 LP014 HDJM Brain; normal pSport 1 LP014 HEFM Adrenal Gland, normal pSport 1 LP014 HBAA Rectum normal pSport 1 LP014 HFDM Rectum tumour pSport 1 LP014 HGAM Colon, normal pSport 1 LP014 HHMM Colon, tumour pSport 1 LP014 HCLB HCLC Human Lung Cancer Lambda Zap II LP015 HRLA L1 Cell line ZAP Express LP015 HHAM Hypothalamus, Alzheimer's pCMVSport 3.0 LP015 HKBA Ku 812F Basophils Line pSport 1 LP015 HS2S Saos2, Dexamethosome Treated pSport 1 LP016 HA5A Lung Carcinoma A549 TNFalpha pSport 1 LP016 activated HTFM TF-1 Cell Line GM-CSF Treated pSport 1 LP016 HYAS Thyroid Tumour pSport 1 LP016 HUTS Larynx Normal pSport 1 LP016 HXOA Larynx Tumor pSport 1 LP016 HEAH Ea.hy.926 cell line pSport 1 LP016 HINA Adenocarcinoma Human pSport 1 LP016 HRMA Lung Mesothelium pSport 1 LP016 HLCL Human Pre-Differentiated Uni-Zap XR LP017 Adipocytes HS2A Saos2 Cells pSport 1 LP020 HS2I Saos2 Cells; Vitamin D3 Treated pSport 1 LP020 HUCM CHME Cell Line, untreated pSport 1 LP020 HEPN Aryepiglottis Normal pSport 1 LP020 HPSN Sinus Piniformis Tumour pSport 1 LP020 HNSA Stomach Normal pSport 1 LP020 HNSM Stomach Tumour pSport 1 LP020 HNLA Liver Normal Met5No pSport 1 LP020 HUTA Liver Tumour Met 5 Tu pSport 1 LP020 HOCN Colon Normal pSport 1 LP020 HOCT Colon Tumor pSport 1 LP020 HTNT Tongue Tumour pSport 1 LP020 HLXN Larynx Normal pSport 1 LP020 HLXT Larynx Tumour pSport 1 LP020 HTYN Thymus pSport 1 LP020 HPLN Placenta pSport 1 LP020 HTNG Tongue Normal pSport 1 LP020 HZAA Thyroid Normal (SDCA2 No) pSport 1 LP020 HWES Thyroid Thyroiditis pSport 1 LP020 HFHD Ficolled Human Stromal Cells, 5Fu pTrip1Ex2 LP021 treated HFHM, HFHN Ficolled Human Stromal Cells, pTrip1Ex2 LP021 Untreated HPCI Hep G2 Cells, lambda library lambda Zap-CMV XR LP021 HBCA, HBCB, HBCC H. Lymph node breast Cancer Uni-ZAP XR LP021 HCOK Chondrocytes pSPORT 1 LP022 HDCA, HDCB, HDCC Dendritic Cells From CD34 Cells pSPORT 1 LP022 HDMA, HDMB CD40 activated monocyte dendritic pSPORT 1 LP022 cells HDDM, HDDN, HDDO LPS activated derived dendritic pSPORT 1 LP022 cells HPCR Hep G2 Cells, PCR library lambda Zap-CMV XR LP022 HAAA, HAAB, HAAC Lung, Cancer (4005313A3): pSPORT 1 LP022 Invasive Poorly Differentiated Lung Adenocarcinoma HIPA, HIPB, HIPC Lung, Cancer (4005163 B7): pSPORT 1 LP022 Invasive, Poorly Diff. Adenocarcinoma, Metastatic HOOH, HOOI Ovary, Cancer: (4004562 B6) pSPORT 1 LP022 Papillary Serous Cystic Neoplasm, Low Malignant Pot HIDA Lung, Normal: (4005313 B1) pSPORT 1 LP022 HUJA, HUJB, HUJC, HUJD, HUJE B-Cells pCMVSport 3.0 LP022 HNOA, HNOB, HNOC, HNOD Ovary, Normal: (9805C040R) pSPORT 1 LP022 HNLM Lung, Normal: (4005313 B1) pSPORT 1 LP022 HSCL Stromal Cells pSPORT 1 LP022 HAAX Lung, Cancer: (4005313 A3) pSPORT 1 LP022 Invasive Poorly-differentiated Metastatic lung adenocarcinoma HUUA, HUUB, HUUC, HUUD B-cells (unstimulated) pTrip1Ex2 LP022 HWWA, HWWB, HWWC, HWWD, B-cells (stimulated) pSPORT 1 LP022 HWWE, HWWF, HWWG HCCC Colon, Cancer: (9808C064R) pCMVSport 3.0 LP023 HPDO HPDP HPDQ HPDR HPD Ovary, Cancer (9809C332): Poorly pSport 1 LP023 differentiated adenocarcinoma HPCO HPCP HPCQ HPCT Ovary, Cancer (15395A1F): Grade pSport 1 LP023 II Papillary Carcinoma HOCM HOCO HOCP HOCQ Ovary, Cancer: (15799A1F) Poorly pSport 1 LP023 differentiated carcinoma HCBM HCBN HCBO Breast, Cancer: (4004943 A5) pSport 1 LP023 HNBT HNBU HNBV Breast, Normal: (4005522B2) pSport 1 LP023 HBCP HBCQ Breast, Cancer: (4005522 A2) pSport 1 LP023 HBCJ Breast, Cancer: (9806C012R) pSport 1 LP023 HSAM HSAN Stromal cells 3.88 pSport 1 LP023 HVCA HVCB HVCC HVCD Ovary, Cancer: (4004332 A2) pSport 1 LP023 HSCK HSEN HSEO Stromal cells (HBM3.18) pSport 1 LP023 HSCP HSCQ stromal cell clone 2.5 pSport 1 LP023 HUXA Breast Cancer: (4005385 A2) pSport 1 LP023 HCOM HCON HCOO HCOP Ovary, Cancer (4004650 A3): Well- pSport 1 LP023 HCOQ Differentiated Micropapillary Serous Carcinoma HBNM Breast, Cancer: (9802C020E) pSport 1 LP023 HVVA HVVB HVVC HVVD Human Bone Marrow, treated pSport 1 LP023 HVVE

Two nonlimiting examples are provided below for isolating a particular clone from the deposited sample of plasmid cDNAs cited for that clone in Table 7. First, a plasmid is directly isolated by screening the clones using a polynucleotide probe corresponding to the nucleotide sequence of SEQ ID NO:X.

Particularly, a specific polynucleotide with 30-40 nucleotides is synthesized using an Applied Biosystems DNA synthesizer according to the sequence reported. The oligonucleotide is labeled, for instance, with ³²P-γ-ATP using T4 polynucleotide kinase and purified according to routine methods. (E.g., Maniatis et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press, Cold Spring, N.Y. (1982)). The plasmid mixture is transformed into a suitable host, as indicated above (such as XL-1 Blue (STRATAGENE™)) using techniques known to those of skill in the art, such as those provided by the vector supplier or in related publications or patents cited above. The transformants are plated on 1.5% agar plates (containing the appropriate selection agent, e.g., ampicillin) to a density of about 150 transformants (colonies) per plate. These plates are screened using Nylon membranes according to routine methods for bacterial colony screening (e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd Edit., (1989), Cold Spring Harbor Laboratory Press, pages 1.93 to 1.104), or other techniques known to those of skill in the art.

Alternatively, two primers of 17-20 nucleotides derived from both ends of the nucleotide sequence of SEQ ID NO:X are synthesized and used to amplify the desired cDNA using the deposited cDNA plasmid as a template. The polymerase chain reaction is carried out under routine conditions, for instance, in 25 μl of reaction mixture with 0.5 μg of the above cDNA template. A convenient reaction mixture is 1.5-5 mM MgCl₂, 0.01% (w/v) gelatin, 20 μM each of dATP, dCTP, dGTP, dTTP, 25 μmol of each primer and 0.25 Unit of Taq polymerase. Thirty five cycles of PCR (denaturation at 94° C. for 1 min; annealing at 55° C. for 1 min; elongation at 72° C. for 1 min) are performed with a Perkin-Elmer Cetus automated thermal cycler. The amplified product is analyzed by agarose gel electrophoresis and the DNA band with expected molecular weight is excised and purified. The PCR product is verified to be the selected sequence by subcloning and sequencing the DNA product.

Several methods are available for the identification of the 5′ or 3′ non-coding portions of a gene which may not be present in the deposited clone. These methods include but are not limited to, filter probing, clone enrichment using specific probes, and protocols similar or identical to 5′ and 3′ “RACE” protocols which are well known in the art. For instance, a method similar to 5′ RACE is available for generating the missing 5′ end of a desired full-length transcript. (Fromont-Racine et al., Nucleic Acids Res. 21(7):1683-1684 (1993)).

Briefly, a specific RNA oligonucleotide is ligated to the 5′ ends of a population of RNA presumably containing full-length gene RNA transcripts. A primer set containing a primer specific to the ligated RNA oligonucleotide and a primer specific to a known sequence of the gene of interest is used to PCR amplify the 5′ portion of the desired full-length gene. This amplified product may then be sequenced and used to generate the full length gene.

This above method starts with total RNA isolated from the desired source, although poly-A+ RNA can be used. The RNA preparation can then be treated with phosphatase if necessary to eliminate 5′ phosphate groups on degraded or damaged RNA which may interfere with the later RNA ligase step. The phosphatase should then be inactivated and the RNA treated with tobacco acid pyrophosphatase in order to remove the cap structure present at the 5′ ends of messenger RNAs. This reaction leaves a 5′ phosphate group at the 5′ end of the cap cleaved RNA which can then be ligated to an RNA oligonucleotide using T4 RNA ligase.

This modified RNA preparation is used as a template for first strand cDNA synthesis using a gene specific oligonucleotide. The first strand synthesis reaction is used as a template for PCR amplification of the desired 5′ end using a primer specific to the ligated RNA oligonucleotide and a primer specific to the known sequence of the gene of interest. The resultant product is then sequenced and analyzed to confirm that the 5′ end sequence belongs to the desired gene.

Example 2 Isolation of Genomic Clones Corresponding to a Polynucleotide

A human genomic P1 library (Genomic Systems, Inc.) is screened by PCR using primers selected for the sequence corresponding to SEQ ID NO:X according to the method described in Example 1. (See also, Sambrook.)

Example 3 Tissue Specific Expression Analysis

The Human Genome Sciences, Inc. (HGS) database is derived from sequencing tissue and/or disease specific cDNA libraries. Libraries generated from a particular tissue are selected and the specific tissue expression pattern of EST groups or assembled contigs within these libraries is determined by comparison of the expression patterns of those groups or contigs within the entire database. ESTs and assembled contigs which show tissue specific expression are selected.

The original clone from which the specific EST sequence was generated, or in the case of an assembled contig, the clone from which the 5′ most EST sequence was generated, is obtained from the catalogued library of clones and the insert amplified by PCR using methods known in the art. The PCR product is denatured and then transferred in 96 or 384 well format to a nylon membrane (Schleicher and Scheull) generating an array filter of tissue specific clones. Housekeeping genes, maize genes, and known tissue specific genes are included on the filters. These targets can be used in signal normalization and to validate assay sensitivity. Additional targets are included to monitor probe length and specificity of hybridization.

Radioactively labeled hybridization probes are generated by first strand cDNA synthesis per the manufacturer's instructions (Life Technologies) from mRNA/RNA samples prepared from the specific tissue being analyzed (e.g., prostate, prostate cancer, ovarian, ovarian cancer, etc.). The hybridization probes are purified by gel exclusion chromatography, quantitated, and hybridized with the array filters in hybridization bottles at 65° C. overnight. The filters are washed under stringent conditions and signals are captured using a Fuji phosphorimager.

Data is extracted using AIS software and following background subtraction, signal normalization is performed. This includes a normalization of filter-wide expression levels between different experimental runs. Genes that are differentially expressed in the tissue of interest are identified.

Example 4 Chromosomal Mapping of the Polynucleotides

An oligonucleotide primer set is designed according to the sequence at the 5′ end of SEQ ID NO:X. This primer preferably spans about 100 nucleotides. This primer set is then used in a polymerase chain reaction under the following set of conditions: 30 seconds, 95° C.; 1 minute, 56° C.; 1 minute, 70° C. This cycle is repeated 32 times followed by one 5 minute cycle at 70° C. Human, mouse, and hamster DNA is used as template in addition to a somatic cell hybrid panel containing individual chromosomes or chromosome fragments (Bios, Inc). The reactions are analyzed on either 8% polyacrylamide gels or 3.5 agarose gels. Chromosome mapping is determined by the presence of an approximately 100 by PCR fragment in the particular somatic cell hybrid.

Example 5 Bacterial Expression of a Polypeptide

A polynucleotide encoding a polypeptide of the present invention is amplified using PCR oligonucleotide primers corresponding to the 5′ and 3′ ends of the DNA sequence, as outlined in Example 1, to synthesize insertion fragments. The primers used to amplify the cDNA insert should preferably contain restriction sites, such as BamHI and XbaI, at the 5′ end of the primers in order to clone the amplified product into the expression vector. For example, BamHI and XbaI correspond to the restriction enzyme sites on the bacterial expression vector pQE-9. (Qiagen, Inc., Chatsworth, Calif.). This plasmid vector encodes antibiotic resistance (Amp^(r)), a bacterial origin of replication (on), an IPTG-regulatable promoter/operator (P/O), a ribosome binding site (RBS), a 6-histidine tag (6-His), and restriction enzyme cloning sites.

The pQE-9 vector is digested with BamHI and XbaI and the amplified fragment is ligated into the pQE-9 vector maintaining the reading frame initiated at the bacterial RBS. The ligation mixture is then used to transform the E. coli strain M15/rep4 (Qiagen, Inc.) which contains multiple copies of the plasmid pREP4, which expresses the lad repressor and also confers kanamycin resistance (Kan^(r)). Transformants are identified by their ability to grow on LB plates and ampicillin/kanamycin resistant colonies are selected. Plasmid DNA is isolated and confirmed by restriction analysis.

Clones containing the desired constructs are grown overnight (0/N) in liquid culture in LB media supplemented with both Amp (100 μg/ml) and Kan (25 μg/ml). The 0/N culture is used to inoculate a large culture at a ratio of 1:100 to 1:250. The cells are grown to an optical density 600 (O.D.⁶⁰⁰) of between 0.4 and 0.6. IPTG (Isopropyl-B-D-thiogalacto pyranoside) is then added to a final concentration of 1 mM. IPTG induces by inactivating the lad repressor, clearing the P/O leading to increased gene expression.

Cells are grown for an extra 3 to 4 hours. Cells are then harvested by centrifugation (20 min at 6000×g). The cell pellet is solubilized in the chaotropic agent 6 Molar Guanidine HCl by stirring for 3-4 hours at 4° C. The cell debris is removed by centrifugation, and the supernatant containing the polypeptide is loaded onto a nickel-nitrilo-tri-acetic acid (“Ni-NTA”) affinity resin column (available from QIAGEN, Inc., supra). Proteins with a 6×His tag bind to the Ni-NTA resin with high affinity and can be purified in a simple one-step procedure (for details see: The QIAexpressionist (1995) QIAGEN, Inc., supra).

Briefly, the supernatant is loaded onto the column in 6 M guanidine-HCl, pH 8. The column is first washed with 10 volumes of 6 M guanidine-HCl, pH 8, then washed with 10 volumes of 6 M guanidine-HCl pH 6, and finally the polypeptide is eluted with 6 M guanidine-HCl, pH 5.

The purified protein is then renatured by dialyzing it against phosphate-buffered saline (PBS) or 50 mM Na-acetate, pH 6 buffer plus 200 mM NaCl. Alternatively, the protein can be successfully refolded while immobilized on the Ni-NTA column. The recommended conditions are as follows: renature using a linear 6M-1M urea gradient in 500 mM NaCl, 20% glycerol, 20 mM Tris/HCl pH 7.4, containing protease inhibitors. The renaturation should be performed over a period of 1.5 hours or more. After renaturation the proteins are eluted by the addition of 250 mM immidazole. Immidazole is removed by a final dialyzing step against PBS or 50 mM sodium acetate pH 6 buffer plus 200 mM NaCl. The purified protein is stored at 4° C. or frozen at −80° C.

In addition to the above expression vector, the present invention further includes an expression vector, called pHE4a (ATCC™ Accession Number 209645, deposited on Feb. 25, 1998) which contains phage operator and promoter elements operatively linked to a polynucleotide of the present invention, called pHE4a. (ATCC™ Accession Number 209645, deposited on Feb. 25, 1998.) This vector contains: 1) a neomycinphosphotransferase gene as a selection marker, 2) an E. coli origin of replication, 3) a T5 phage promoter sequence, 4) two lac operator sequences, 5) a Shine-Delgarno sequence, and 6) the lactose operon repressor gene (lacIq). The origin of replication (oriC) is derived from pUC19 (LTI, Gaithersburg, Md.). The promoter and operator sequences are made synthetically.

DNA can be inserted into the pHE4a by restricting the vector with NdeI and XbaI, BamHI, XhoI, or Asp718, running the restricted product on a gel, and isolating the larger fragment (the stuffer fragment should be about 310 base pairs). The DNA insert is generated according to the PCR protocol described in Example 1, using PCR primers having restriction sites for NdeI (5′ primer) and XbaI, BamHI, XhoI, or Asp718 (3′ primer). The PCR insert is gel purified and restricted with compatible enzymes. The insert and vector are ligated according to standard protocols.

The engineered vector could easily be substituted in the above protocol to express protein in a bacterial system.

Example 6 Purification of a Polypeptide from an Inclusion Body

The following alternative method can be used to purify a polypeptide expressed in E coli when it is present in the form of inclusion bodies. Unless otherwise specified, all of the following steps are conducted at 4-10° C.

Upon completion of the production phase of the E. coli fermentation, the cell culture is cooled to 4-10° C. and the cells harvested by continuous centrifugation at 15,000 rpm (Heraeus Sepatech). On the basis of the expected yield of protein per unit weight of cell paste and the amount of purified protein required, an appropriate amount of cell paste, by weight, is suspended in a buffer solution containing 100 mM Tris, 50 mM EDTA, pH 7.4. The cells are dispersed to a homogeneous suspension using a high shear mixer.

The cells are then lysed by passing the solution through a microfluidizer (Microfluidics, Corp. or APV Gaulin, Inc.) twice at 4000-6000 psi. The homogenate is then mixed with NaCl solution to a final concentration of 0.5 M NaCl, followed by centrifugation at 7000×g for 15 min. The resultant pellet is washed again using 0.5M NaCl, 100 mM Tris, 50 mM EDTA, pH 7.4.

The resulting washed inclusion bodies are solubilized with 1.5 M guanidine hydrochloride (GuHCl) for 2-4 hours. After 7000×g centrifugation for 15 min., the pellet is discarded and the polypeptide containing supernatant is incubated at 4° C. overnight to allow further GuHCl extraction.

Following high speed centrifugation (30,000×g) to remove insoluble particles, the GuHCl solubilized protein is refolded by quickly mixing the GuHCl extract with 20 volumes of buffer containing 50 mM sodium, pH 4.5, 150 mM NaCl, 2 mM EDTA by vigorous stirring. The refolded diluted protein solution is kept at 4° C. without mixing for 12 hours prior to further purification steps.

To clarify the refolded polypeptide solution, a previously prepared tangential filtration unit equipped with 0.16 μm membrane filter with appropriate surface area (e.g., Filtron), equilibrated with 40 mM sodium acetate, pH 6.0 is employed. The filtered sample is loaded onto a cation exchange resin (e.g., Poros HS-50, Perseptive Biosystems). The column is washed with 40 mM sodium acetate, pH 6.0 and eluted with 250 mM, 500 mM, 1000 mM, and 1500 mM NaCl in the same buffer, in a stepwise manner. The absorbance at 280 nm of the effluent is continuously monitored. Fractions are collected and further analyzed by SDS-PAGE.

Fractions containing the polypeptide are then pooled and mixed with 4 volumes of water. The diluted sample is then loaded onto a previously prepared set of tandem columns of strong anion (Poros HQ-50, Perseptive Biosystems) and weak anion (Poros CM-20, Perseptive Biosystems) exchange resins. The columns are equilibrated with 40 mM sodium acetate, pH 6.0. Both columns are washed with 40 mM sodium acetate, pH 6.0, 200 mM NaCl. The CM-20 column is then eluted using a 10 column volume linear gradient ranging from 0.2 M NaCl, 50 mM sodium acetate, pH 6.0 to 1.0 M NaCl, 50 mM sodium acetate, pH 6.5. Fractions are collected under constant A₂₈₀ monitoring of the effluent. Fractions containing the polypeptide (determined, for instance, by 16% SDS-PAGE) are then pooled.

The resultant polypeptide should exhibit greater than 95% purity after the above refolding and purification steps. No major contaminant bands should be observed from Commassie blue stained 16% SDS-PAGE gel when 5 μg of purified protein is loaded. The purified protein can also be tested for endotoxin/LPS contamination, and typically the LPS content is less than 0.1 ng/ml according to LAL assays.

Example 7 Cloning and Expression of a Polypeptide in a Baculovirus Expression System

In this example, the plasmid shuttle vector pA2 is used to insert a polynucleotide into a baculovirus to express a polypeptide. This expression vector contains the strong polyhedrin promoter of the Autographa californica nuclear polyhedrosis virus (AcMNPV) followed by convenient restriction sites such as BamHI, Xba I and Asp718. The polyadenylation site of the simian virus 40 (“SV40”) is used for efficient polyadenylation. For easy selection of recombinant virus, the plasmid contains the beta-galactosidase gene from E. coli under control of a weak Drosophila promoter in the same orientation, followed by the polyadenylation signal of the polyhedrin gene. The inserted genes are flanked on both sides by viral sequences for cell-mediated homologous recombination with wild-type viral DNA to generate a viable virus that express the cloned polynucleotide.

Many other baculovirus vectors can be used in place of the vector above, such as pAc373, pVL941, and pAcIM1, as one skilled in the art would readily appreciate, as long as the construct provides appropriately located signals for transcription, translation, secretion and the like, including a signal peptide and an in-frame AUG as required. Such vectors are described, for instance, in Luckow et al., Virology 170:31-39 (1989).

Specifically, the cDNA sequence contained in the deposited clone, including the AUG initiation codon, is amplified using the PCR protocol described in Example 1. If a naturally occurring signal sequence is used to produce the polypeptide of the present invention, the pA2 vector does not need a second signal peptide. Alternatively, the vector can be modified (pA2 GP) to include a baculovirus leader sequence, using the standard methods described in Summers et al., “A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures,” Texas Agricultural Experimental Station Bulletin No. 1555 (1987).

The amplified fragment is isolated from a 1% agarose gel using a commercially available kit (“GENECLEAN™,” BIO 101 Inc., La Jolla, Calif.). The fragment then is digested with appropriate restriction enzymes and again purified on a 1% agarose gel.

The plasmid is digested with the corresponding restriction enzymes and optionally, can be dephosphorylated using calf intestinal phosphatase, using routine procedures known in the art. The DNA is then isolated from a 1% agarose gel using a commercially available kit (“GENECLEAN™” BIO 101 Inc., La Jolla, Calif.).

The fragment and the dephosphorylated plasmid are ligated together with T4 DNA ligase. E. coli HB101 or other suitable E. coli hosts such as XL-1 Blue (STRATAGENE™ Cloning Systems, La Jolla, Calif.) cells are transformed with the ligation mixture and spread on culture plates. Bacteria containing the plasmid are identified by digesting DNA from individual colonies and analyzing the digestion product by gel electrophoresis. The sequence of the cloned fragment is confirmed by DNA sequencing.

Five μg of a plasmid containing the polynucleotide is co-transfected with 1.0 μg of a commercially available linearized baculovirus DNA (“BACULOGOLD™ baculovirus DNA, Pharmingen, San Diego, Calif.), using the lipofection method described by Feigner et al., Proc. Natl. Acad. Sci. USA 84:7413-7417 (1987). One μg of BACULOGOLD™ virus DNA and 5 μg of the plasmid are mixed in a sterile well of a microtiter plate containing 50 μl of serum-free Grace's medium (Life Technologies Inc., Gaithersburg, Md.). Afterwards, 10 μA LIPOFECTIN™ plus 90 μl Grace's medium are added, mixed and incubated for 15 minutes at room temperature. Then the transfection mixture is added drop-wise to 519 insect cells (ATCC™ CRL 1711) seeded in a 35 mm tissue culture plate with 1 ml Grace's medium without serum. The plate is then incubated for 5 hours at 27° C. The transfection solution is then removed from the plate and 1 ml of Grace's insect medium supplemented with 10% fetal calf serum is added. Cultivation is then continued at 27° C. for four days.

After four days the supernatant is collected and a plaque assay is performed, as described by Summers and Smith, supra. An agarose gel with “Blue Gal” (Life Technologies Inc., Gaithersburg) is used to allow easy identification and isolation of gal-expressing clones, which produce blue-stained plaques. (A detailed description of a “plaque assay” of this type can also be found in the user's guide for insect cell culture and baculovirology distributed by Life Technologies Inc., Gaithersburg, page 9-10.) After appropriate incubation, blue stained plaques are picked with the tip of a micropipettor (e.g., Eppendorf). The agar containing the recombinant viruses is then resuspended in a microcentrifuge tube containing 200 μl of Grace's medium and the suspension containing the recombinant baculovirus is used to infect 519 cells seeded in 35 mm dishes. Four days later the supernatants of these culture dishes are harvested and then they are stored at 4° C.

To verify the expression of the polypeptide, Sf9 cells are grown in Grace's medium supplemented with 10% heat-inactivated FBS. The cells are infected with the recombinant baculovirus containing the polynucleotide at a multiplicity of infection (“MOI”) of about 2. If radiolabeled proteins are desired, 6 hours later the medium is removed and is replaced with SF900 II medium minus methionine and cysteine (available from Life Technologies Inc., Rockville, Md.). After 42 hours, 5 μCi of ³⁵S-methionine and 5 μCi ³⁵S-cysteine (available from Amersham) are added. The cells are further incubated for 16 hours and then are harvested by centrifugation. The proteins in the supernatant as well as the intracellular proteins are analyzed by SDS-PAGE followed by autoradiography (if radiolabeled).

Microsequencing of the amino acid sequence of the amino terminus of purified protein may be used to determine the amino terminal sequence of the produced protein.

Example 8 Expression of a Polypeptide in Mammalian Cells

The polypeptide of the present invention can be expressed in a mammalian cell. A typical mammalian expression vector contains a promoter element, which mediates the initiation of transcription of mRNA, a protein coding sequence, and signals required for the termination of transcription and polyadenylation of the transcript. Additional elements include enhancers, Kozak sequences and intervening sequences flanked by donor and acceptor sites for RNA splicing. Highly efficient transcription is achieved with the early and late promoters from SV40, the long terminal repeats (LTRs) from Retroviruses, e.g., RSV, HTLVI, HIVI and the early promoter of the cytomegalovirus (CMV). However, cellular elements can also be used (e.g., the human actin promoter).

Suitable expression vectors for use in practicing the present invention include, for example, vectors such as pSVL and pMSG (PHARMACIA™, Uppsala, Sweden), pRSVcat (ATCC™ 37152), pSV2dhfr (ATCC™ 37146), pBC12MI (ATCC™ 67109), pCMVSport 2.0, and pCMVSport 3.0. Mammalian host cells that could be used include, human Hela, 293, H9 and Jurkat cells, mouse NIH3T3 and C127 cells, Cos 1, Cos 7 and CV1, quail QC1-3 cells, mouse L cells and Chinese hamster ovary (CHO) cells.

Alternatively, the polypeptide can be expressed in stable cell lines containing the polynucleotide integrated into a chromosome. The co-transfection with a selectable marker such as DHFR, gpt, neomycin, or hygromycin allows the identification and isolation of the transfected cells.

The transfected gene can also be amplified to express large amounts of the encoded protein. The DHFR (dihydrofolate reductase) marker is useful in developing cell lines that carry several hundred or even several thousand copies of the gene of interest. (See, e.g., Alt, F. W., et al., J. Biol. Chem. 253:1357-1370 (1978); Hamlin, J. L. and Ma, C., Biochem. et Biophys. Acta, 1097:107-143 (1990); Page, M. J. and Sydenham, M. A., Biotechnology 9:64-68 (1991)). Another useful selection marker is the enzyme glutamine synthase (GS) (Murphy et al., Biochem J. 227:277-279 (1991); Bebbington et al., Bio/Technology 10:169-175 (1992). Using these markers, the mammalian cells are grown in selective medium and the cells with the highest resistance are selected. These cell lines contain the amplified gene(s) integrated into a chromosome. Chinese hamster ovary (CHO) and NSO cells are often used for the production of proteins.

Derivatives of the plasmid pSV2-dhfr (ATCC™ Accession No. 37146), the expression vectors pC4 (ATCC™ Accession No. 209646) and pC6 (ATCC™ Accession No. 209647) contain the strong promoter (LTR) of the Rous Sarcoma Virus (Cullen et al., Molecular and Cellular Biology, 438-447 (March, 1985)) plus a fragment of the CMV-enhancer (Boshart et al., Cell 41:521-530 (1985)). Multiple cloning sites, e.g., with the restriction enzyme cleavage sites BamHI, XbaI and Asp718, facilitate the cloning of the gene of interest. The vectors also contain the 3′ intron, the polyadenylation and termination signal of the rat preproinsulin gene, and the mouse DHFR gene under control of the SV40 early promoter.

Specifically, the plasmid pC6, for example, is digested with appropriate restriction enzymes and then dephosphorylated using calf intestinal phosphates by procedures known in the art. The vector is then isolated from a 1% agarose gel.

A polynucleotide of the present invention is amplified according to the protocol outlined in Example 1. If a naturally occurring signal sequence is used to produce the polypeptide of the present invention, the vector does not need a second signal peptide. Alternatively, if a naturally occurring signal sequence is not used, the vector can be modified to include a heterologous signal sequence. (See, e.g., International Publication No. WO 96/34891.)

The amplified fragment is isolated from a 1% agarose gel using a commercially available kit (“GENECLEAN™,” BIO 101 Inc., La Jolla, Calif.). The fragment then is digested with appropriate restriction enzymes and again purified on a 1% agarose gel.

The amplified fragment is then digested with the same restriction enzyme and purified on a 1% agarose gel. The isolated fragment and the dephosphorylated vector are then ligated with T4 DNA ligase. E. coli HB101 or XL-1 Blue cells are then transformed and bacteria are identified that contain the fragment inserted into plasmid pC6 using, for instance, restriction enzyme analysis.

Chinese hamster ovary cells lacking an active DHFR gene is used for transfection. Five μg of the expression plasmid pC6 or pC4 is cotransfected with 0.5 μg of the plasmid pSVneo using LIPOFECTIN™ (Felgner et al., supra). The plasmid pSV2-neo contains a dominant selectable marker, the neo gene from Tn5 encoding an enzyme that confers resistance to a group of antibiotics including G418. The cells are seeded in alpha minus MEM supplemented with 1 mg/ml G418. After 2 days, the cells are trypsinized and seeded in hybridoma cloning plates (Greiner, Germany) in alpha minus MEM supplemented with 10, 25, or 50 ng/ml of methotrexate plus 1 mg/ml G418. After about 10-14 days single clones are trypsinized and then seeded in 6-well petri dishes or 10 ml flasks using different concentrations of methotrexate (50 nM, 100 nM, 200 nM, 400 nM, 800 nM). Clones growing at the highest concentrations of methotrexate are then transferred to new 6-well plates containing even higher concentrations of methotrexate (1 μM, 2 μM, 5 μM, 10 mM, 20 mM). The same procedure is repeated until clones are obtained which grow at a concentration of 100-200 μM. Expression of the desired gene product is analyzed, for instance, by SDS-PAGE and Western blot or by reversed phase HPLC analysis.

Example 9 Protein Fusions

The polypeptides of the present invention are preferably fused to other proteins. These fusion proteins can be used for a variety of applications. For example, fusion of the present polypeptides to His-tag, HA-tag, protein A, IgG domains, and maltose binding protein facilitates purification. (See Example 5; see also EP A 394,827; Traunecker, et al., Nature 331:84-86 (1988)). Similarly, fusion to IgG-1, IgG-3, and albumin increases the halflife time in vivo. Nuclear localization signals fused to the polypeptides of the present invention can target the protein to a specific subcellular localization, while covalent heterodimer or homodimers can increase or decrease the activity of a fusion protein. Fusion proteins can also create chimeric molecules having more than one function. Finally, fusion proteins can increase solubility and/or stability of the fused protein compared to the non-fused protein. All of the types of fusion proteins described above can be made by modifying the following protocol, which outlines the fusion of a polypeptide to an IgG molecule, or the protocol described in Example 5.

Briefly, the human Fc portion of the IgG molecule can be PCR amplified, using primers that span the 5′ and 3′ ends of the sequence described below. These primers also should have convenient restriction enzyme sites that will facilitate cloning into an expression vector, preferably a mammalian expression vector.

For example, if pC4 (ATCC™ Accession No. 209646) is used, the human Fc portion can be ligated into the BamHI cloning site. Note that the 3′ BamHI site should be destroyed. Next, the vector containing the human Fc portion is re-restricted with BamHI, linearizing the vector, and a polynucleotide of the present invention, isolated by the PCR protocol described in Example 1, is ligated into this BamHI site. Note that the polynucleotide is cloned without a stop codon, otherwise a fusion protein will not be produced.

If the naturally occurring signal sequence is used to produce the polypeptide of the present invention, pC4 does not need a second signal peptide. Alternatively, if the naturally occurring signal sequence is not used, the vector can be modified to include a heterologous signal sequence. (See, e.g., International Publication No. WO 96/34891.)

Human IgG Fc region: (SEQ ID NO: 1) GGGATCCGGAGCCCAAATCTTCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAATTCGAG GGTGCACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACTCCTGA GGTCACATGCGTGGTGGTGGACGTAAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGAC GGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTG GTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCA ACAAAGCCCTCCCAACCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACA GGTGTACACCCTGCCCCCATCCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCA AAGGCTTCTATCCAAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAA GACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGA GCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACG CAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGAGTGCGACGGCCGCGACTCTAGAGGAT

Example 10 Production of an Antibody from a Polypeptide a) Hybridoma Technology

The antibodies of the present invention can be prepared by a variety of methods. (See, Current Protocols, Chapter 2.) As one example of such methods, cells expressing a polypeptide of the present invention are administered to an animal to induce the production of sera containing polyclonal antibodies. In a preferred method, a preparation of a polypeptide of the present invention is prepared and purified to render it substantially free of natural contaminants. Such a preparation is then introduced into an animal in order to produce polyclonal antisera of greater specific activity.

Monoclonal antibodies specific for a polypeptide of the present invention are prepared using hybridoma technology (Kohler et al., Nature 256:495 (1975); Kohler et al., Eur. J. Immunol. 6:511 (1976); Kohler et al., Eur. J. Immunol. 6:292 (1976); Hammerling et al., in: Monoclonal Antibodies and T-Cell Hybridomas, Elsevier, N.Y., pp. 563-681 (1981)). In general, an animal (preferably a mouse) is immunized with a polypeptide of the present invention or, more preferably, with a secreted polypeptide-expressing cell. Such polypeptide-expressing cells are cultured in any suitable tissue culture medium, preferably in Earle's modified Eagle's medium supplemented with 10% fetal bovine serum (inactivated at about 56° C.), and supplemented with about 10 g/l of nonessential amino acids, about 1,000 U/ml of penicillin, and about 100 μg/ml of streptomycin.

The splenocytes of such mice are extracted and fused with a suitable myeloma cell line. Any suitable myeloma cell line may be employed in accordance with the present invention; however, it is preferable to employ the parent myeloma cell line (SP2O), available from the ATCC™. After fusion, the resulting hybridoma cells are selectively maintained in HAT medium, and then cloned by limiting dilution as described by Wands et al. (Gastroenterology 80:225-232 (1981)). The hybridoma cells obtained through such a selection are then assayed to identify clones which secrete antibodies capable of binding the polypeptide of the present invention.

Alternatively, additional antibodies capable of binding to a polypeptide of the present invention can be produced in a two-step procedure using anti-idiotypic antibodies. Such a method makes use of the fact that antibodies are themselves antigens, and therefore, it is possible to obtain an antibody that binds to a second antibody. In accordance with this method, protein specific antibodies are used to immunize an animal, preferably a mouse. The splenocytes of such an animal are then used to produce hybridoma cells, and the hybridoma cells are screened to identify clones which produce an antibody whose ability to bind to the polypeptide-specific antibody can be blocked by said polypeptide. Such antibodies comprise anti-idiotypic antibodies to the polypeptide-specific antibody and are used to immunize an animal to induce formation of further polypeptide-specific antibodies.

For in vivo use of antibodies in humans, an antibody is “humanized”. Such antibodies can be produced using genetic constructs derived from hybridoma cells producing the monoclonal antibodies described above. Methods for producing chimeric and humanized antibodies are known in the art and are discussed herein. (See, for review, Morrison, Science 229:1202 (1985); Oi et al., BioTechniques 4:214 (1986); Cabilly et al., U.S. Pat. No. 4,816,567; Taniguchi et al., EP 171496; Morrison et al., EP 173494; Neuberger et al., WO 8601533; Robinson et al., International Publication No. WO 8702671; Boulianne et al., Nature 312:643 (1984); Neuberger et al., Nature 314:268 (1985)).

b) Isolation of Antibody Fragments Directed Against a Polypeptide of the Present Invention from a Library of scFvs

Naturally occurring V-genes isolated from human PBLs are constructed into a library of antibody fragments which contain reactivities against a polypeptide of the present invention to which the donor may or may not have been exposed (see e.g., U.S. Pat. No. 5,885,793 incorporated herein by reference in its entirety).

Rescue of the Library. A library of scFvs is constructed from the RNA of human PBLs as described in International Publication No. WO 92/01047. To rescue phage displaying antibody fragments, approximately 10⁹ E. coli harboring the phagemid are used to inoculate 50 ml of 2×TY containing 1% glucose and 100 μg/ml of ampicillin (2×TY-AMP-GLU) and grown to an O.D. of 0.8 with shaking. Five ml of this culture is used to inoculate 50 ml of 2×TY-AMP-GLU, 2×108 TU of delta gene 3 helper (M13 delta gene III, see International Publication No. WO 92/01047) are added and the culture incubated at 37° C. for 45 minutes without shaking and then at 37° C. for 45 minutes with shaking. The culture is centrifuged at 4000 r.p.m. for 10 min. and the pellet resuspended in 2 liters of 2×TY containing 100 μg/ml ampicillin and 50 μg/ml kanamycin and grown overnight. Phage are prepared as described in International Publication No. WO 92/01047.

M13 delta gene III is prepared as follows: M13 delta gene III helper phage does not encode gene III protein, hence the phage(mid) displaying antibody fragments have a greater avidity of binding to antigen. Infectious M13 delta gene III particles are made by growing the helper phage in cells harboring a pUC19 derivative supplying the wild type gene III protein during phage morphogenesis. The culture is incubated for 1 hour at 37° C. without shaking and then for a further hour at 37° C. with shaking. Cells are spun down (IEC-Centra 8,400 r.p.m. for 10 min), resuspended in 300 ml 2×TY broth containing 100 μg ampicillin/ml and 25 μg kanamycin/ml (2×TY-AMP-KAN) and grown overnight, shaking at 37° C. Phage particles are purified and concentrated from the culture medium by two PEG-precipitations (Sambrook et al., 1990), resuspended in 2 ml PBS and passed through a 0.45 μm filter (Minisart NML; Sartorius) to give a final concentration of approximately 10¹³ transducing units/ml (ampicillin-resistant clones).

Panning of the Library Immunotubes (Nunc) are coated overnight in PBS with 4 ml of either 100 μg/ml or 10 μg/ml of a polypeptide of the present invention. Tubes are blocked with 2% Marvel-PBS for 2 hours at 37° C. and then washed 3 times in PBS. Approximately 10¹³ TU of phage is applied to the tube and incubated for 30 minutes at room temperature tumbling on an over and under turntable and then left to stand for another 1.5 hours. Tubes are washed 10 times with PBS 0.1% Tween-20 and 10 times with PBS. Phage are eluted by adding 1 ml of 100 mM triethylamine and rotating 15 minutes on an under and over turntable after which the solution is immediately neutralized with 0.5 ml of 1.0M Tris-HCl, pH 7.4. Phage are then used to infect 10 ml of mid-log E. coli TG1 by incubating eluted phage with bacteria for 30 minutes at 37° C. The E. coli are then plated on TYE plates containing 1% glucose and 100 μg/ml ampicillin. The resulting bacterial library is then rescued with delta gene 3 helper phage as described above to prepare phage for a subsequent round of selection. This process is then repeated for a total of 4 rounds of affinity purification with tube-washing increased to 20 times with PBS, 0.1% Tween-20 and 20 times with PBS for rounds 3 and 4.

Characterization of Binders. Eluted phage from the 3rd and 4th rounds of selection are used to infect E. coli HB 2151 and soluble scFv is produced (Marks, et al., 1991) from single colonies for assay. ELISAs are performed with microtitre plates coated with either 10 pg/ml of the polypeptide of the present invention in 50 mM bicarbonate pH 9.6. Clones positive in ELISA are further characterized by PCR fingerprinting (see, e.g., International Publication No. WO 92/01047) and then by sequencing. These ELISA positive clones may also be further characterized by techniques known in the art, such as, for example, epitope mapping, binding affinity, receptor signal transduction, ability to block or competitively inhibit antibody/antigen binding, and competitive agonistic or antagonistic activity.

Example 11 Method of Determining Alterations in a Gene Corresponding to a Polynucleotide

RNA isolated from entire families or individual patients presenting with an immune disease or disorder is isolated. cDNA is then generated from these RNA samples using protocols known in the art. (See, Sambrook.) The cDNA is then used as a template for PCR, employing primers surrounding regions of interest in SEQ ID NO:X; and/or the nucleotide sequence of the cDNA contained in ATCC™ Deposit No:Z. Suggested PCR conditions consist of 35 cycles at 95 degrees C. for 30 seconds; 60-120 seconds at 52-58 degrees C.; and 60-120 seconds at 70 degrees C., using buffer solutions described in Sidransky et al., Science 252:706 (1991).

PCR products are then sequenced using primers labeled at their 5′ end with T4 polynucleotide kinase, employing SequiTherm Polymerase (Epicentre Technologies). The intron-exon boundaries of selected exons is also determined and genomic PCR products analyzed to confirm the results. PCR products harboring suspected mutations are then cloned and sequenced to validate the results of the direct sequencing.

PCR products are cloned into T-tailed vectors as described in Holton et al., Nucleic Acids Research, 19:1156 (1991) and sequenced with T7 polymerase (United States Biochemical). Affected individuals are identified by mutations not present in unaffected individuals.

Genomic rearrangements are also observed as a method of determining alterations in a gene corresponding to a polynucleotide. Genomic clones isolated according to Example 2 are nick-translated with digoxigenindeoxy-uridine 5′-triphosphate (Boehringer Manheim), and FISH performed as described in Johnson et al., Methods Cell Biol. 35:73-99 (1991). Hybridization with the labeled probe is carried out using a vast excess of human cot-1 DNA for specific hybridization to the corresponding genomic locus.

Chromosomes are counterstained with 4,6-diamino-2-phenylidole and propidium iodide, producing a combination of C- and R-bands. Aligned images for precise mapping are obtained using a triple-band filter set (Chroma Technology, Brattleboro, Vt.) in combination with a cooled charge-coupled device camera (Photometrics, Tucson, Ariz.) and variable excitation wavelength filters. (Johnson et al., Genet. Anal. Tech. Appl., 8:75 (1991)). Image collection, analysis and chromosomal fractional length measurements are performed using the ISee Graphical Program System. (Inovision Corporation, Durham, N.C.) Chromosome alterations of the genomic region hybridized by the probe are identified as insertions, deletions, and translocations. These alterations are used as a diagnostic marker for an associated disease.

Example 12 Method of Detecting Abnormal Levels of a Polypeptide in a Biological Sample

A polypeptide of the present invention can be detected in a biological sample, and if an increased or decreased level of the polypeptide is detected, this polypeptide is a marker for a particular phenotype. Methods of detection are numerous, and thus, it is understood that one skilled in the art can modify the following assay to fit their particular needs.

For example, antibody-sandwich ELISAs are used to detect polypeptides in a sample, preferably a biological sample. Wells of a microtiter plate are coated with specific antibodies, at a final concentration of 0.2 to 10 μg/ml. The antibodies are either monoclonal or polyclonal and are produced by the method described in Example 10. The wells are blocked so that non-specific binding of the polypeptide to the well is reduced.

The coated wells are then incubated for >2 hours at RT with a sample containing the polypeptide. Preferably, serial dilutions of the sample should be used to validate results. The plates are then washed three times with deionized or distilled water to remove unbound polypeptide.

Next, 50 μl of specific antibody-alkaline phosphatase conjugate, at a concentration of 25-400 ng, is added and incubated for 2 hours at room temperature. The plates are again washed three times with deionized or distilled water to remove unbound conjugate.

Add 75 μl of 4-methylumbelliferyl phosphate (MUP) or p-nitrophenyl phosphate (NPP) substrate solution to each well and incubate 1 hour at room temperature. Measure the reaction by a microtiter plate reader. Prepare a standard curve, using serial dilutions of a control sample, and plot polypeptide concentration on the X-axis (log scale) and fluorescence or absorbance of the Y-axis (linear scale). Interpolate the concentration of the polypeptide in the sample using the standard curve.

Example 13 Formulation

The invention also provides methods of preventing, treating and/or ameliorating an immune disease or disorder by administration to a subject of an effective amount of a Therapeutic. By therapeutic is meant polynucleotides or polypeptides of the invention (including fragments and variants), agonists or antagonists thereof, and/or antibodies thereto, in combination with a pharmaceutically acceptable carrier type (e.g., a sterile carrier).

The Therapeutic will be formulated and dosed in a fashion consistent with good medical practice, taking into account the clinical condition of the individual patient (especially the side effects of treatment with the Therapeutic alone), the site of delivery, the method of administration, the scheduling of administration, and other factors known to practitioners. The “effective amount” for purposes herein is thus determined by such considerations.

As a general proposition, the total pharmaceutically effective amount of the Therapeutic administered parenterally per dose will be in the range of about 1 ug/kg/day to 10 mg/kg/day of patient body weight, although, as noted above, this will be subject to therapeutic discretion. More preferably, this dose is at least 0.01 mg/kg/day, and most preferably for humans between about 0.01 and 1 mg/kg/day for the hormone. If given continuously, the Therapeutic is typically administered at a dose rate of about 1 μg/kg/hour to about 50 μg/kg/hour, either by 1-4 injections per day or by continuous subcutaneous infusions, for example, using a mini-pump. An intravenous bag solution may also be employed. The length of treatment needed to observe changes and the interval following treatment for responses to occur appears to vary depending on the desired effect.

Therapeutics can be are administered orally, rectally, parenterally, intracistemally, intravaginally, intraperitoneally, topically (as by powders, ointments, gels, drops or transdermal patch), bucally, or as an oral or nasal spray. “Pharmaceutically acceptable carrier” refers to a non-toxic solid, semisolid or liquid filler, diluent, encapsulating material or formulation auxiliary of any. The term “parenteral” as used herein refers to modes of administration which include intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous and intraarticular injection and infusion.

Therapeutics of the invention are also suitably administered by sustained-release systems. Suitable examples of sustained-release Therapeutics are administered orally, rectally, parenterally, intracistemally, intravaginally, intraperitoneally, topically (as by powders, ointments, gels, drops or transdermal patch), bucally, or as an oral or nasal spray. “Pharmaceutically acceptable carrier” refers to a non-toxic solid, semisolid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. The term “parenteral” as used herein refers to modes of administration which include intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous and intraarticular injection and infusion.

Therapeutics of the invention are also suitably administered by sustained-release systems. Suitable examples of sustained-release Therapeutics include suitable polymeric materials (such as, for example, semi-permeable polymer matrices in the form of shaped articles, e.g., films, or mirocapsules), suitable hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, and sparingly soluble derivatives (such as, for example, a sparingly soluble salt).

Sustained-release matrices include polylactides (U.S. Pat. No. 3,773,919, EP 58,481), copolymers of L-glutamic acid and gamma-ethyl-L-glutamate (Sidman et al., Biopolymers 22:547-556 (1983)), poly (2-hydroxyethyl methacrylate) (Langer et al., J. Biomed. Mater. Res. 15:167-277 (1981), and Langer, Chem. Tech. 12:98-105 (1982)), ethylene vinyl acetate (Langer et al., Id.) or poly-D-(−)-3-hydroxybutyric acid (EP 133,988).

In a preferred embodiment, polypeptide, polynucleotide, and antibody compositions of the invention are formulated in a biodegradable, polymeric drug delivery system, for example as described in U.S. Pat. Nos. 4,938,763; 5,278,201; 5,278,202; 5,324,519; 5,340,849; and 5,487,897 and in International Publication Numbers WO01/35929, WO00/24374, and WO00/06117 which are hereby incorporated by reference in their entirety. In specific preferred embodiments the polypeptide, polynucleotide, and antibody compositions of the invention are formulated using the ATRIGEL® Biodegradable System of Atrix Laboratories, Inc. (Fort Collins, Colorado).

Examples of biodegradable polymers which can be used in the formulation of polypeptide, polynucleotide, and antibody compositions, include but are not limited to, polylactides, polyglycolides, polycaprolactones, polyanhydrides, polyamides, polyurethanes, polyesteramides, polyorthoesters, polydioxanones, polyacetals, polyketals, polycarbonates, polyorthocarbonates, polyphosphazenes, polyhydroxybutyrates, polyhydroxyvalerates, polyalkylene oxalates, polyalkylene succinates, poly(malic acid), poly(amino acids), poly(methyl vinyl ether), poly(maleic anhydride), polyvinylpyrrolidone, polyethylene glycol, polyhydroxycellulose, chitin, chitosan, and copolymers, terpolymers, or combinations or mixtures of the above materials. The preferred polymers are those that have a lower degree of crystallization and are more hydrophobic. These polymers and copolymers are more soluble in the biocompatible solvents than the highly crystalline polymers such as polyglycolide and chitin which also have a high degree of hydrogen-bonding. Preferred materials with the desired solubility parameters are the polylactides, polycaprolactones, and copolymers of these with glycolide in which there are more amorphous regions to enhance solubility. In specific preferred embodiments, the biodegradable polymers which can be used in the formulation of polypeptide, polynucleotide, and antibody compositions are poly(lactide-co-glycolides). Polymer properties such as molecular weight, hydrophobicity, and lactide/glycolide ratio may be modified to obtain the desired polypeptide, polynucleotide, or antibody release profile (See, e.g., Ravivarapu et al., Journal of Pharmaceutical Sciences 89:732-741 (2000), which is hereby incorporated by reference in its entirety).

It is also preferred that the solvent for the biodegradable polymer be non-toxic, water miscible, and otherwise biocompatible. Examples of such solvents include, but are not limited to, N-methyl-2-pyrrolidone, 2-pyrrolidone, C2 to C6 alkanols, C1 to C15 alchohols, dils, triols, and tetraols such as ethanol, glycerine propylene glycol, butanol; C3 to C15 alkyl ketones such as acetone, diethyl ketone and methyl ethyl ketone; C3 to C15 esters such as methyl acetate, ethyl acetate, ethyl lactate; alkyl ketones such as methyl ethyl ketone, C1 to C15 amides such as dimethylformamide, dimethylacetamide and caprolactam; C3 to C20 ethers such as tetrahydrofuran, or solketal; tweens, triacetin, propylene carbonate, decylmethylsulfoxide, dimethyl sulfoxide, oleic acid, 1-dodecylazacycloheptan-2-one, Other preferred solvents are benzyl alchohol, benzyl benzoate, dipropylene glycol, tributyrin, ethyl oleate, glycerin, glycofural, isopropyl myristate, isopropyl palmitate, oleic acid, polyethylene glycol, propylene carbonate, and triethyl citrate. The most preferred solvents are N-methyl-2-pyrrolidone, 2-pyrrolidone, dimethyl sulfoxide, triacetin, and propylene carbonate because of the solvating ability and their compatibility.

Additionally, formulations comprising polypeptide, polynucleotide, and antibody compositions and a biodegradable polymer may also include release-rate modification agents and/or pore-forming agents. Examples of release-rate modification agents include, but are not limited to, fatty acids, triglycerides, other like hydrophobic compounds, organic solvents, plasticizing compounds and hydrophilic compounds. Suitable release rate modification agents include, for example, esters of mono-, di-, and tricarboxylic acids, such as 2-ethoxyethyl acetate, methyl acetate, ethyl acetate, diethyl phthalate, dimethyl phthalate, dibutyl phthalate, dimethyl adipate, dimethyl succinate, dimethyl oxalate, dimethyl citrate, triethyl citrate, acetyl tributyl citrate, acetyl triethyl citrate, glycerol triacetate, di(n-butyl) sebecate, and the like; polyhydroxy alcohols, such as propylene glycol, polyethylene glycol, glycerin, sorbitol, and the like; fatty acids; triesters of glycerol, such as triglycerides, epoxidized soybean oil, and other epoxidized vegetable oils; sterols, such as cholesterol; alcohols, such as C.sub.6-C.sub.12 alkanols, 2-ethoxyethanol. The release rate modification agent may be used singly or in combination with other such agents. Suitable combinations of release rate modification agents include, but are not limited to, glycerin/propylene glycol, sorbitol/glycerine, ethylene oxide/propylene oxide, butylene glycol/adipic acid, and the like. Preferred release rate modification agents include, but are not limited to, dimethyl citrate, triethyl citrate, ethyl heptanoate, glycerin, and hexanediol. Suitable pore-forming agents that may be used in the polymer composition include, but are not limited to, sugars such as sucrose and dextrose, salts such as sodium chloride and sodium carbonate, polymers such as hydroxylpropylcellulose, carboxymethylcellulose, polyethylene glycol, and polyvinylpyrrolidone. Solid crystals that will provide a defined pore size, such as salt or sugar, are preferred.

In specific preferred embodiments the polypeptide, polynucleotide, and antibody compositions of the invention are formulated using the BEMA™ BioErodible Mucoadhesive System, MCA™ MucoCutaneous Absorption System, SMP™ Solvent MicroParticle System, or BCP™ BioCompatible Polymer System of Atrix Laboratories, Inc. (Fort Collins, Colo.).

Sustained-release Therapeutics also include liposomally entrapped Therapeutics of the invention (see generally, Langer, Science 249:1527-1533 (1990); Treat et al., in Liposomes in the Therapy of Infectious Disease and Cancer, Lopez-Berestein and Fidler (eds.), Liss, New York, pp. 317-327 and 353-365 (1989)). Liposomes containing the Therapeutic are prepared by methods known per se: DE 3,218,121; Epstein et al., Proc. Natl. Acad. Sci. (USA) 82:3688-3692 (1985); Hwang et al., Proc. Natl. Acad. Sci. (USA) 77:4030-4034 (1980); EP 52,322; EP 36,676; EP 88,046; EP 143,949; EP 142,641; Japanese Pat. Appl. 83-118008; U.S. Pat. Nos. 4,485,045 and 4,544,545; and EP 102,324. Ordinarily, the liposomes are of the small (about 200-800 Angstroms) unilamellar type in which the lipid content is greater than about 30 mol. percent cholesterol, the selected proportion being adjusted for the optimal Therapeutic.

In yet an additional embodiment, the Therapeutics of the invention are delivered by way of a pump (see Langer, supra; Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987); Buchwald et al., Surgery 88:507 (1980); Saudek et al., N. Engl. J. Med. 321:574 (1989)).

Other controlled release systems are discussed in the review by Langer (Science 249:1527-1533 (1990)).

For parenteral administration, in one embodiment, the Therapeutic is formulated generally by mixing it at the desired degree of purity, in a unit dosage injectable form (solution, suspension, or emulsion), with a pharmaceutically acceptable carrier, i.e., one that is non-toxic to recipients at the dosages and concentrations employed and is compatible with other ingredients of the formulation. For example, the formulation preferably does not include oxidizing agents and other compounds that are known to be deleterious to the Therapeutic.

Generally, the formulations are prepared by contacting the Therapeutic uniformly and intimately with liquid carriers or finely divided solid carriers or both. Then, if necessary, the product is shaped into the desired formulation. Preferably the carrier is a parenteral carrier, more preferably a solution that is isotonic with the blood of the recipient. Examples of such carrier vehicles include water, saline, Ringer's solution, and dextrose solution. Non-aqueous vehicles such as fixed oils and ethyl oleate are also useful herein, as well as liposomes.

The carrier suitably contains minor amounts of additives such as substances that enhance isotonicity and chemical stability. Such materials are non-toxic to recipients at the dosages and concentrations employed, and include buffers such as phosphate, citrate, succinate, acetic acid, and other organic acids or their salts; antioxidants such as ascorbic acid; low molecular weight (less than about ten residues) polypeptides, e.g., polyarginine or tripeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids, such as glycine, glutamic acid, aspartic acid, or arginine; monosaccharides, disaccharides, and other carbohydrates including cellulose or its derivatives, glucose, manose, or dextrins; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; counterions such as sodium; and/or nonionic surfactants such as polysorbates, poloxamers, or PEG.

The Therapeutic is typically formulated in such vehicles at a concentration of about 0.1 mg/ml to 100 mg/ml, preferably 1-10 mg/ml, at a pH of about 3 to 8. It will be understood that the use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of polypeptide salts.

Any pharmaceutical used for therapeutic administration can be sterile. Sterility is readily accomplished by filtration through sterile filtration membranes (e.g., 0.2 micron membranes). Therapeutics generally are placed into a container having a sterile access port, for example, an intravenous solution bag or vial having a stopper pierceable by a hypodermic injection needle.

Therapeutics ordinarily will be stored in unit or multi-dose containers, for example, sealed ampoules or vials, as an aqueous solution or as a lyophilized formulation for reconstitution. As an example of a lyophilized formulation, 10-ml vials are filled with 5 ml of sterile-filtered 1% (w/v) aqueous Therapeutic solution, and the resulting mixture is lyophilized. The infusion solution is prepared by reconstituting the lyophilized Therapeutic using bacteriostatic Water-for-Injection.

The invention also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the Therapeutics of the invention. Associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration. In addition, the Therapeutics may be employed in conjunction with other therapeutic compounds.

The Therapeutics of the invention may be administered alone or in combination with adjuvants. Adjuvants that may be administered with the Therapeutics of the invention include, but are not limited to, alum, alum plus deoxycholate (ImmunoAg), MTP-PE (Biocine Corp.), QS21 (Genentech, Inc.), BCG (e.g., THERACYS®), MPL and nonviable prepartions of Corynebacterium parvum. In a specific embodiment, Therapeutics of the invention are administered in combination with alum. In another specific embodiment, Therapeutics of the invention are administered in combination with QS-21. Further adjuvants that may be administered with the Therapeutics of the invention include, but are not limited to, Monophosphoryl lipid immunomodulator, AdjuVax 100a, QS-21, QS-18, CRL1005, Aluminum salts, MF-59, and Virosomal adjuvant technology. Vaccines that may be administered with the Therapeutics of the invention include, but are not limited to, vaccines directed toward protection against MMR (measles, mumps, rubella), polio, varicella, tetanus/diptheria, hepatitis A, hepatitis B, haemophilus influenzae B, whooping cough, pneumonia, influenza, Lyme's Disease, rotavirus, cholera, yellow fever, Japanese encephalitis, poliomyelitis, rabies, typhoid fever, and pertussis. Combinations may be administered either concomitantly, e.g., as an admixture, separately but simultaneously or concurrently; or sequentially. This includes presentations in which the combined agents are administered together as a therapeutic mixture, and also procedures in which the combined agents are administered separately but simultaneously, e.g., as through separate intravenous lines into the same individual. Administration “in combination” further includes the separate administration of one of the compounds or agents given first, followed by the second.

The Therapeutics of the invention may be administered alone or in combination with other therapeutic agents. Therapeutic agents that may be administered in combination with the Therapeutics of the invention, include but not limited to, chemotherapeutic agents, antibiotics, steroidal and non-steroidal anti-inflammatories, conventional immunotherapeutic agents, and/or therapeutic treatments described below. Combinations may be administered either concomitantly, e.g., as an admixture, separately but simultaneously or concurrently; or sequentially. This includes presentations in which the combined agents are administered together as a therapeutic mixture, and also procedures in which the combined agents are administered separately but simultaneously, e.g., as through separate intravenous lines into the same individual. Administration “in combination” further includes the separate administration of one of the compounds or agents given first, followed by the second.

In one embodiment, the Therapeutics of the invention are administered in combination with an anticoagulant. Anticoagulants that may be administered with the compositions of the invention include, but are not limited to, heparin, low molecular weight heparin, warfarin sodium (e.g., COUMADIN®), dicumarol, 4-hydroxycoumarin, anisindione (e.g., MIRADON™), acenocoumarol (e.g., nicoumalone, SINTHROME™), indan-1,3-dione, phenprocoumon (e.g., MARCUMAR™), ethyl biscoumacetate (e.g., TROMEXAN™), and aspirin. In a specific embodiment, compositions of the invention are administered in combination with heparin and/or warfarin. In another specific embodiment, compositions of the invention are administered in combination with warfarin. In another specific embodiment, compositions of the invention are administered in combination with warfarin and aspirin. In another specific embodiment, compositions of the invention are administered in combination with heparin. In another specific embodiment, compositions of the invention are administered in combination with heparin and aspirin.

In another embodiment, the Therapeutics of the invention are administered in combination with thrombolytic drugs. Thrombolytic drugs that may be administered with the compositions of the invention include, but are not limited to, plasminogen, lys-plasminogen, alpha2-antiplasmin, streptokinae (e.g., KABIKINASE™) antiresplace (e.g., EMINASE™), tissue plasminogen activator (t-PA, altevase, ACTIVASE™), urokinase (e.g., ABBOKINASE™), sauruplase, (Prourokinase, single chain urokinase), and aminocaproic acid (e.g., AMICAR™). In a specific embodiment, compositions of the invention are administered in combination with tissue plasminogen activator and aspirin.

In another embodiment, the Therapeutics of the invention are administered in combination with antiplatelet drugs. Antiplatelet drugs that may be administered with the compositions of the invention include, but are not limited to, aspirin, dipyridamole (e.g., PERSANTINE™), and ticlopidine (e.g., TICLID™).

In specific embodiments, the use of anti-coagulants, thrombolytic and/or antiplatelet drugs in combination with Therapeutics of the invention is contemplated for the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of thrombosis, arterial thrombosis, venous thrombosis, thromboembolism, pulmonary embolism, atherosclerosis, myocardial infarction, transient ischemic attack, unstable angina. In specific embodiments, the use of anticoagulants, thrombolytic drugs and/or antiplatelet drugs in combination with Therapeutics of the invention is contemplated for the prevention of occulsion of saphenous grafts, for reducing the risk of periprocedural thrombosis as might accompany angioplasty procedures, for reducing the risk of stroke in patients with atrial fibrillation including nonrheumatic atrial fibrillation, for reducing the risk of embolism associated with mechanical heart valves and or mitral valves disease. Other uses for the therapeutics of the invention, alone or in combination with antiplatelet, anticoagulant, and/or thrombolytic drugs, include, but are not limited to, the prevention of occlusions in extracorporeal devices (e.g., intravascular canulas, vascular access shunts in hemodialysis patients, hemodialysis machines, and cardiopulmonary bypass machines).

In certain embodiments, Therapeutics of the invention are administered in combination with antiretroviral agents, nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), and/or protease inhibitors (PIs). NRTIs that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, RETROVIR™ (zidovudine/AZT), VIDEX™ (didanosine/ddI), HIVID™ (zalcitabine/ddC), ZERIT™ (stavudine/d4T), EPIVIR™ (lamivudine/3TC), and COMBIVIR™ (zidovudine/lamivudine). NNRTIs that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, VIRAMUNE™ (nevirapine), RESCRIPTOR™ (delavirdine), and SUSTIVA™ (efavirenz). Protease inhibitors that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, CRIXIVAN™ (indinavir), NORVIR™ (ritonavir), INVIRASE™ (saquinavir), and VIRACEPT™ (nelfinavir). In a specific embodiment, antiretroviral agents, nucleoside reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors, and/or protease inhibitors may be used in any combination with Therapeutics of the invention to treat AIDS and/or to prevent or treat HIV infection.

Additional NRTIs include LODENOSINE™ (F-ddA; an acid-stable adenosine NRTI; Triangle/Abbott; COVIRACIL™ (emtricitabine/FTC; structurally related to lamivudine (3TC) but with 3- to 10-fold greater activity in vitro; Triangle/Abbott); dOTC (BCH-10652, also structurally related to lamivudine but retains activity against a substantial proportion of lamivudine-resistant isolates; Biochem Pharma); Adefovir (refused approval for anti-HIV therapy by FDA; Gilead Sciences); PREVEON® (Adefovir Dipivoxil, the active prodrug of adefovir; its active form is PMEA-pp); TENOFOVIR™ (bis-POC PMPA, a PMPA prodrug; Gilead); DAPD/DXG (active metabolite of DAPD; Triangle/Abbott); D-D4FC (related to 3TC, with activity against AZT/3TC-resistant virus); GW420867X (Glaxo Wellcome); ZIAGEN™ (abacavir/159U89; Glaxo Wellcome Inc.); CS-87 (3′ azido-2′,3′-dideoxyuridine; WO 99/66936); and S-acyl-2-thioethyl (SATE)-bearing prodrug forms of β-L-FD4C and β-L-FddC (WO 98/17281).

Additional NNRTIs include COACTINON™ (Emivirine/MKC-442, potent NNRTI of the HEPT class; Triangle/Abbott); CAPRAVIRINE™ (AG-1549/S-1153, a next generation NNRTI with activity against viruses containing the K103N mutation; Agouron); PNU-142721 (has 20- to 50-fold greater activity than its predecessor delavirdine and is active against K103N mutants; PHARMACIA™ & Upjohn); DPC-961 and DPC-963 (second-generation derivatives of efavirenz, designed to be active against viruses with the K103N mutation; DUPONT™); GW-420867×(has 25-fold greater activity than HBY097 and is active against K103N mutants; Glaxo Wellcome); CALANOLIDE A (naturally occurring agent from the latex tree; active against viruses containing either or both the Y181C and K103N mutations); and Propolis (WO 99/49830).

Additional protease inhibitors include LOPINAVIR™ (ABT378/r; Abbott Laboratories); BMS-232632 (an azapeptide; Bristol-Myres Squibb); TIPRANAVIR™ (PNU-140690, a non-peptic dihydropyrone; PHARMACIA™ & Upjohn); PD-178390 (a nonpeptidic dihydropyrone; Parke-Davis); BMS 232632 (an azapeptide; Bristol-Myers Squibb); L-756,423 (an indinavir analog; Merck); DMP-450 (a cyclic urea compound; Avid & DUPONT™); AG-1776 (a peptidomimetic with in vitro activity against protease inhibitor-resistant viruses; Agouron); VX-175/GW-433908 (phosphate prodrug of amprenavir; Vertex & Glaxo Welcome); CGP61755 (Ciba); and AGENERASE™ (amprenavir; Glaxo Wellcome Inc.).

Additional antiretroviral agents include fusion inhibitors/gp41 binders. Fusion inhibitors/gp41 binders include T-20 (a peptide from residues 643-678 of the HIV gp41 transmembrane protein ectodomain which binds to gp41 in its resting state and prevents transformation to the fusogenic state; Trimeris) and T-1249 (a second-generation fusion inhibitor; Trimeris).

Additional antiretroviral agents include fusion inhibitors/chemokine receptor antagonists. Fusion inhibitors/chemokine receptor antagonists include CXCR4 antagonists such as AMD 3100 (a bicyclam), SDF-1 and its analogs, and ALX40-4C (a cationic peptide), T22 (an 18 amino acid peptide; Trimeris) and the T22 analogs T134 and T140; CCR5 antagonists such as RANTES (9-68), AOP-RANTES, NNY-RANTES, and TAK-779; and CCR5/CXCR4 antagonists such as NSC 651016 (a distamycin analog). Also included are CCR2B, CCR3, and CCR6 antagonists. Chemokine recpetor agonists such as RANTES, SDF-1, MIP-1α, MIP-1β, etc., may also inhibit fusion.

Additional antiretroviral agents include integrase inhibitors. Integrase inhibitors include dicaffeoylquinic (DFQA) acids; L-chicoric acid (a dicaffeoyltartaric (DCTA) acid); quinalizarin (QLC) and related anthraquinones; ZINTEVIR™ (AR 177, an oligonucleotide that probably acts at cell surface rather than being a true integrase inhibitor; Arondex); and naphthols such as those disclosed in WO 98/50347.

Additional antiretroviral agents include hydroxyurea-like compounds such as BCX-34 (a purine nucleoside phosphorylase inhibitor; Biocryst); ribonucleotide reductase inhibitors such as DIDOX™ (Molecules for Health); inosine monophosphate dehydrogenase (IMPDH) inhibitors sucha as VX-497 (Vertex); and mycopholic acids such as CellCept (mycophenolate mofetil; Roche).

Additional antiretroviral agents include inhibitors of viral integrase, inhibitors of viral genome nuclear translocation such as arylene bis(methylketone) compounds; inhibitors of HW entry such as AOP-RANTES, NNY-RANTES, RANTES-IgG fusion protein, soluble complexes of RANTES and glycosaminoglycans (GAG), and AMD-3100; nucleocapsid zinc finger inhibitors such as dithiane compounds; targets of HIV Tat and Rev; and pharmacoenhancers such as ABT-378.

Other antiretroviral therapies and adjunct therapies include cytokines and lymphokines such as MIP-1α, MIP-1β, SDF-1α, IL-2, PROLEUKIN™ (aldesleukin/L2-7001; Chiron), IL-4, IL-10, IL-12, and IL-13; interferons such as IFN-α2a; antagonists of TNFs, NFκB, GM-CSF, M-CSF, and IL-10; agents that modulate immune activation such as cyclosporin and prednisone; vaccines such as Remune™ (HIV Immunogen), APL 400-003 (Apollon), recombinant gp120 and fragments, bivalent (B/E) recombinant envelope glycoprotein, rgp120CM235, MN rgp120, SF-2 rgp120, gp120/soluble CD4 complex, Delta JR-FL protein, branched synthetic peptide derived from discontinuous gp120 C3/C4 domain, fusion-competent immunogens, and Gag, Pol, Nef, and Tat vaccines; gene-based therapies such as genetic suppressor elements (GSEs; WO 98/54366), and intrakines (genetically modified CC chemokines targetted to the ER to block surface expression of newly synthesized CCR5 (Yang et al., PNAS 94:11567-72 (1997); Chen et al., Nat. Med. 3:1110-16 (1997)); antibodies such as the anti-CXCR4 antibody 12G5, the anti-CCR5 antibodies 2D7, 5C7, PA8, PA9, PA10, PA11, PAl2, and PA14, the anti-CD4 antibodies Q4120 and RPA-T4, the anti-CCR3 antibody 7B11, the anti-gp120 antibodies 17b, 48d, 447-52D, 257-D, 268-D and 50.1, anti-Tat antibodies, anti-TNF-a antibodies, and monoclonal antibody 33A; aryl hydrocarbon (AH) receptor agonists and antagonists such as TCDD, 3,3′,4,4′,5-pentachlorobiphenyl, 3,3′,4,4′-tetrachlorobiphenyl, and α-naphthoflavone (WO 98/30213); and antioxidants such as γ-L-glutamyl-L-cysteine ethyl ester (γ-GCE; WO 99/56764).

In a further embodiment, the Therapeutics of the invention are administered in combination with an antiviral agent. Antiviral agents that may be administered with the Therapeutics of the invention include, but are not limited to, acyclovir, ribavirin, amantadine, and remantidine.

In other embodiments, Therapeutics of the invention may be administered in combination with anti-opportunistic infection agents. Anti-opportunistic agents that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, TRIMETHOPRIM-SULFAMETHOXAZOLE™, DAPSONE™, PENTAMIDINE™, ATOVAQUONE™, ISONIAZID™, RIFAMPIN™, PYRAZINAMIDE™, ETHAMBUTOL™, RIFABUTIN™, CLARITHROMYCIN™, AZITHROMYCIN™, GANCICLOVIR™, FOSCARNET™, CIDOFOVIR™, FLUCONAZOLE™, ITRACONAZOLE™, KETOCONAZOLE™, ACYCLOVIR™, FAMCICOLVIR™, PYRIMETHAMINE™, LEUCOVORIN™, NEUPOGEN™ (filgrastim/G-CSF), and LEUKINE™ (sargramostim/GM-CSF). In a specific embodiment, Therapeutics of the invention are used in any combination with TRIMETHOPRIM-SULFAMETHOXAZOLE™, DAPSONE™, PENTAMIDINE™, and/or ATOVAQUONE™ to prophylactically treat or prevent an opportunistic Pneumocystis carinii pneumonia infection. In another specific embodiment, Therapeutics of the invention are used in any combination with ISONIAZID™ RIFAMPIN™, PYRAZINAMIDE™, and/or ETHAMBUTOL™ to prophylactically treat or prevent an opportunistic Mycobacterium avium complex infection. In another specific embodiment, Therapeutics of the invention are used in any combination with RIFABUTIN™, CLARITHROMYCIN™, and/or AZITHROMYCIN™ to prophylactically treat or prevent an opportunistic Mycobacterium tuberculosis infection. In another specific embodiment, Therapeutics of the invention are used in any combination with GANCICLOVIR™, FOSCARNET™, and/or CIDOFOVIR™ to prophylactically treat or prevent an opportunistic cytomegalovirus infection. In another specific embodiment, Therapeutics of the invention are used in any combination with FLUCONAZOLE™ ITRACONAZOLE™, and/or KETOCONAZOLE™ to prophylactically treat or prevent an opportunistic fungal infection. In another specific embodiment, Therapeutics of the invention are used in any combination with ACYCLOVIR™ and/or FAMCICOLVIR™ to prophylactically treat or prevent an opportunistic herpes simplex virus type I and/or type II infection. In another specific embodiment, Therapeutics of the invention are used in any combination with PYRIMETHAMINE™ and/or LEUCOVORIN™ to prophylactically treat or prevent an opportunistic Toxoplasma gondii infection. In another specific embodiment, Therapeutics of the invention are used in any combination with LEUCOVORIN™ and/or NEUPOGEN™ to prophylactically treat or prevent an opportunistic bacterial infection.

In a further embodiment, the Therapeutics of the invention are administered in combination with an antibiotic agent. Antibiotic agents that may be administered with the Therapeutics of the invention include, but are not limited to, amoxicillin, beta-lactamases, aminoglycosides, beta-lactam (glycopeptide), beta-lactamases, Clindamycin, chloramphenicol, cephalosporins, ciprofloxacin, erythromycin, fluoroquinolones, macrolides, metronidazole, penicillins, quinolones, rapamycin, rifampin, streptomycin, sulfonamide, tetracyclines, trimethoprim, trimethoprim-sulfamethoxazole, and vancomycin.

In other embodiments, the Therapeutics of the invention are administered in combination with immunestimulants Immunostimulants that may be administered in combination with the Therapeutics of the invention include, but are not limited to, levamisole (e.g., ERGAMISOL™), isoprinosine (e.g. INOSIPLEXT™), interferons (e.g. interferon alpha), and interleukins (e.g., IL-2).

In other embodiments, Therapeutics of the invention are administered in combination with immunosuppressive agents. Immunosuppressive agents that may be administered in combination with the Therapeutics of the invention include, but are not limited to, steroids, cyclosporine, cyclosporine analogs, cyclophosphamide methylprednisone, prednisone, azathioprine, FK-506, 15-deoxyspergualin, and other immunosuppressive agents that act by suppressing the function of responding T cells. Other immunosuppressive agents that may be administered in combination with the Therapeutics of the invention include, but are not limited to, prednisolone, methotrexate, thalidomide, methoxsalen, rapamycin, leflunomide, mizoribine (BREDININ™), brequinar, deoxyspergualin, and azaspirane (SKF 105685), ORTHOCLONE OKT® 3 (muromonab-CD3), SANDIMMUNE™, NEORAL™, SANGDYA™ (cyclosporine), PROGRAF® (FK506, tacrolimus), CELLCEPT® (mycophenolate motefil, of which the active metabolite is mycophenolic acid), IMURAN™ (azathioprine), glucocorticosteroids, adrenocortical steroids such as DELTASONE™ (prednisone) and HYDELTRASOL™ (prednisolone), FOLEX™ and MEXATE™ (methotrxate), OXSORALEN-ULTRA™ (methoxsalen) and RAPAMUNE™ (sirolimus). In a specific embodiment, immunosuppressants may be used to prevent rejection of organ or bone marrow transplantation.

In an additional embodiment, Therapeutics of the invention are administered alone or in combination with one or more intravenous immune globulin preparations. Intravenous immune globulin preparations that may be administered with the Therapeutics of the invention include, but not limited to, GAMMAR™, IVEEGAM™, SANDOGLOBULIN™, GAMMAGARD S/D™, ATGAM™ (antithymocyte glubulin), and GAMIMUNE™. In a specific embodiment, Therapeutics of the invention are administered in combination with intravenous immune globulin preparations in transplantation therapy (e.g., bone marrow transplant).

In certain embodiments, the Therapeutics of the invention are administered alone or in combination with an anti-inflammatory agent. Anti-inflammatory agents that may be administered with the Therapeutics of the invention include, but are not limited to, corticosteroids (e.g. betamethasone, budesonide, cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisolone, prednisone, and triamcinolone), nonsteroidal anti-inflammatory drugs (e.g., diclofenac, diflunisal, etodolac, fenoprofen, floctafenine, flurbiprofen, ibuprofen, indomethacin, ketoprofen, meclofenamate, mefenamic acid, meloxicam, nabumetone, naproxen, oxaprozin, phenylbutazone, piroxicam, sulindac, tenoxicam, tiaprofenic acid, and tolmetin.), as well as antihistamines, aminoarylcarboxylic acid derivatives, arylacetic acid derivatives, arylbutyric acid derivatives, arylcarboxylic acids, arylpropionic acid derivatives, pyrazoles, pyrazolones, salicylic acid derivatives, thiazinecarboxamides, e-acetamidocaproic acid, S-adenosylmethionine, 3-amino-4-hydroxybutyric acid, amixetrine, bendazac, benzydamine, bucolome, difenpiramide, ditazol, emorfazone, guaiazulene, nabumetone, nimesulide, orgotein, oxaceprol, paranyline, perisoxal, pifoxime, proquazone, proxazole, and tenidap.

In an additional embodiment, the compositions of the invention are administered alone or in combination with an anti-angiogenic agent. Anti-angiogenic agents that may be administered with the compositions of the invention include, but are not limited to, Angiostatin (Entremed, Rockville, Md.), Troponin-1 (Boston Life Sciences, Boston, Mass.), anti-Invasive Factor, retinoic acid and derivatives thereof, paclitaxel (TAXOL™), Suramin, Tissue Inhibitor of Metalloproteinase-1, Tissue Inhibitor of Metalloproteinase-2, VEGI, Plasminogen Activator Inhibitor-1, Plasminogen Activator Inhibitor-2, and various forms of the lighter “d group” transition metals.

Lighter “d group” transition metals include, for example, vanadium, molybdenum, tungsten, titanium, niobium, and tantalum species. Such transition metal species may form transition metal complexes. Suitable complexes of the above-mentioned transition metal species include oxo transition metal complexes.

Representative examples of vanadium complexes include oxo vanadium complexes such as vanadate and vanadyl complexes. Suitable vanadate complexes include metavanadate and orthovanadate complexes such as, for example, ammonium metavanadate, sodium metavanadate, and sodium orthovanadate. Suitable vanadyl complexes include, for example, vanadyl acetylacetonate and vanadyl sulfate including vanadyl sulfate hydrates such as vanadyl sulfate mono- and trihydrates.

Representative examples of tungsten and molybdenum complexes also include oxo complexes. Suitable oxo tungsten complexes include tungstate and tungsten oxide complexes. Suitable tungstate complexes include ammonium tungstate, calcium tungstate, sodium tungstate dihydrate, and tungstic acid. Suitable tungsten oxides include tungsten (IV) oxide and tungsten (VI) oxide. Suitable oxo molybdenum complexes include molybdate, molybdenum oxide, and molybdenyl complexes. Suitable molybdate complexes include ammonium molybdate and its hydrates, sodium molybdate and its hydrates, and potassium molybdate and its hydrates. Suitable molybdenum oxides include molybdenum (VI) oxide, molybdenum (VI) oxide, and molybdic acid. Suitable molybdenyl complexes include, for example, molybdenyl acetylacetonate. Other suitable tungsten and molybdenum complexes include hydroxo derivatives derived from, for example, glycerol, tartaric acid, and sugars.

A wide variety of other anti-angiogenic factors may also be utilized within the context of the present invention. Representative examples include, but are not limited to, platelet factor 4; protamine sulphate; sulphated chitin derivatives (prepared from queen crab shells), (Murata et al., Cancer Res. 51:22-26, (1991)); Sulphated Polysaccharide Peptidoglycan Complex (SP-PG) (the function of this compound may be enhanced by the presence of steroids such as estrogen, and tamoxifen citrate); Staurosporine; modulators of matrix metabolism, including for example, proline analogs, cishydroxyproline, d,L-3,4-dehydroproline, Thiaproline, alpha,alpha-dipyridyl, aminopropionitrile fumarate; 4-propyl-5-(4-pyridinyl)-2(3H)-oxazolone; Methotrexate; Mitoxantrone; Heparin; Interferons; 2 Macroglobulin-serum; ChIMP-3 (Pavloff et al., J. Bio. Chem. 267:17321-17326, (1992)); Chymostatin (Tomkinson et al., Biochem J. 286:475-480, (1992)); Cyclodextrin Tetradecasulfate; Eponemycin; Camptothecin; Fumagillin (Ingber et al., Nature 348:555-557, (1990)); Gold Sodium Thiomalate (“GST”; Matsubara and Ziff, J. Clin. Invest. 79:1440-1446, (1987)); anticollagenase-serum; alpha2-antiplasmin (Holmes et al., J. Biol. Chem. 262(4):1659-1664, (1987)); Bisantrene (National Cancer Institute); Lobenzarit disodium (N-(2)-carboxyphenyl-4-chloroanthronilic acid disodium or “CCA”; (Takeuchi et al., Agents Actions 36:312-316, (1992)); and metalloproteinase inhibitors such as BB94.

Additional anti-angiogenic factors that may also be utilized within the context of the present invention include Thalidomide, (Celgene, Warren, N.J.); Angiostatic steroid; AGM-1470 (H. Brem and J. Folkman J Pediatr. Surg. 28:445-51 (1993)); an integrin alpha v beta 3 antagonist (C. Storgard et al., J Clin. Invest. 103:47-54 (1999)); carboxynaminolmidazole; Carboxyamidotriazole (CAI) (National Cancer Institute, Bethesda, Md.); Conbretastatin A-4 (CA4P) (OXiGENE, Boston, Mass.); Squalamine (Magainin Pharmaceuticals, Plymouth Meeting, Pa.); TNP-470, (Tap Pharmaceuticals, Deerfield, Ill.); ZD-0101 AstraZeneca (London, UK); APRA (CT2584); Benefin, Byrostatin-1 (SC339555); CGP-41251 (PKC 412); CM101; Dexrazoxane (ICRF 187); DMXAA; Endostatin; Flavopridiol; Genestein; GTE; ImmTher; Iressa (ZD1839); Octreotide (Somatostatin); Panretin; Penacillamine; Photopoint; PI-88; Prinomastat (AG-3340) Purlytin; Suradista (FCE26644); Tamoxifen (Nolvadex); Tazarotene; Tetrathiomolybdate; Xeloda (Capecitabine); and 5-Fluorouracil.

Anti-angiogenic agents that may be administed in combination with the compounds of the invention may work through a variety of mechanisms including, but not limited to, inhibiting proteolysis of the extracellular matrix, blocking the function of endothelial cell-extracellular matrix adhesion molecules, by antagonizing the function of angiogenesis inducers such as growth factors, and inhibiting integrin receptors expressed on proliferating endothelial cells. Examples of anti-angiogenic inhibitors that interfere with extracellular matrix proteolysis and which may be administered in combination with the compositions of the invention include, but are not limited to, AG-3340 (Agouron, La Jolla, Calif.), BAY-12-9566 (Bayer, West Haven, Conn.), BMS-275291 (Bristol Myers Squibb, Princeton, N.J.), CGS-27032A (Novartis, East Hanover, N.J.), Marimastat (British Biotech, Oxford, UK), and Metastat (Aeterna, St-Foy, Quebec). Examples of anti-angiogenic inhibitors that act by blocking the function of endothelial cell-extracellular matrix adhesion molecules and which may be administered in combination with the compositions of the invention include, but are not limited to, EMD-121974 (Merck KcgaA Darmstadt, Germany) and Vitaxin (Ixsys, La Jolla, Calif./Medimmune, Gaithersburg, Md.). Examples of anti-angiogenic agents that act by directly antagonizing or inhibiting angiogenesis inducers and which may be administered in combination with the compositions of the invention include, but are not limited to, Angiozyme (Ribozyme, Boulder, Colo.), Anti-VEGF antibody (Genentech, S. San Francisco, Calif.), PTK-787/ZK-225846 (Novartis, Basel, Switzerland), SU-101 (Sugen, S. San Francisco, Calif.), SU-5416 (Sugen/PHARMACIA™ Upjohn, Bridgewater, N.J.), and SU-6668 (Sugen). Other anti-angiogenic agents act to indirectly inhibit angiogenesis. Examples of indirect inhibitors of angiogenesis which may be administered in combination with the compositions of the invention include, but are not limited to, IM-862 (Cytran, Kirkland, Wash.), Interferon-alpha, IL-12 (Roche, Nutley, N.J.), and Pentosan polysulfate (Georgetown University, Washington, D.C.).

In particular embodiments, the use of compositions of the invention in combination with anti-angiogenic agents is contemplated for the treatment, prevention, and/or amelioration of an autoimmune disease, such as for example, an autoimmune disease described herein.

In a particular embodiment, the use of compositions of the invention in combination with anti-angiogenic agents is contemplated for the treatment, prevention, and/or amelioration of arthritis. In a more particular embodiment, the use of compositions of the invention in combination with anti-angiogenic agents is contemplated for the treatment, prevention, and/or amelioration of rheumatoid arthritis.

In another embodiment, the polynucleotides encoding a polypeptide of the present invention are administered in combination with an angiogenic protein, or polynucleotides encoding an angiogenic protein. Examples of angiogenic proteins that may be administered with the compositions of the invention include, but are not limited to, acidic and basic fibroblast growth factors, VEGF-1, VEGF-2, VEGF-3, epidermal growth factor alpha and beta, platelet-derived endothelial cell growth factor, platelet-derived growth factor, tumor necrosis factor alpha, hepatocyte growth factor, insulin-like growth factor, colony stimulating factor, macrophage colony stimulating factor, granulocyte/macrophage colony stimulating factor, and nitric oxide synthase.

In additional embodiments, compositions of the invention are administered in combination with a chemotherapeutic agent. Chemotherapeutic agents that may be administered with the Therapeutics of the invention include, but are not limited to alkylating agents such as nitrogen mustards (for example, Mechlorethamine, cyclophosphamide, Cyclophosphamide Ifosfamide, Melphalan (L-sarcolysin), and Chlorambucil), ethylenimines and methylmelamines (for example, Hexamethylmelamine and Thiotepa), alkyl sulfonates (for example, Busulfan), nitrosoureas (for example, Carmustine (BCNU), Lomustine (CCNU), Semustine (methyl-CCNU), and Streptozocin (streptozotocin)), triazenes (for example, Dacarbazine (DTIC; dimethyltriazenoimidazolecarboxamide)), folic acid analogs (for example, Methotrexate (amethopterin)), pyrimidine analogs (for example, Fluorouacil (5-fluorouracil; 5-FU), Floxuridine (fluorodeoxyuridine; FudR), and Cytarabine (cytosine arabinoside)), purine analogs and related inhibitors (for example, Mercaptopurine (6-mercaptopurine; 6-MP), Thioguanine (6-thioguanine; TG), and Pentostatin (2′-deoxycoformycin)), vinca alkaloids (for example, Vinblastine (VLB, vinblastine sulfate)) and Vincristine (vincristine sulfate)), epipodophyllotoxins (for example, Etoposide and Teniposide), antibiotics (for example, Dactinomycin (actinomycin D), Daunorubicin (daunomycin; rubidomycin), Doxorubicin, Bleomycin, Plicamycin (mithramycin), and Mitomycin (mitomycin C), enzymes (for example, L-Asparaginase), biological response modifiers (for example, Interferon-alpha and interferon-alpha-2b), platinum coordination compounds (for example, Cisplatin (cis-DDP) and Carboplatin), anthracenedione (Mitoxantrone), substituted ureas (for example, Hydroxyurea), methylhydrazine derivatives (for example, Procarbazine (N-methylhydrazine; MIH), adrenocorticosteroids (for example, Prednisone), progestins (for example, Hydroxyprogesterone caproate, Medroxyprogesterone, Medroxyprogesterone acetate, and Megestrol acetate), estrogens (for example, Diethylstilbestrol (DES), Diethylstilbestrol diphosphate, Estradiol, and Ethinyl estradiol), antiestrogens (for example, Tamoxifen), androgens (Testosterone proprionate, and Fluoxymesterone), antiandrogens (for example, Flutamide), gonadotropin-releasing horomone analogs (for example, Leuprolide), other hormones and hormone analogs (for example, methyltestosterone, estramustine, estramustine phosphate sodium, chlorotrianisene, and testolactone), and others (for example, dicarbazine, glutamic acid, and mitotane).

In one embodiment, the compositions of the invention are administered in combination with one or more of the following drugs: infliximab (also known as Remicade™ Centocor, Inc.), Trocade (Roche, RO-32-3555), Leflunomide (also known as Arava™ from Hoechst Marion Roussel), Kineret™ (an IL-1 Receptor antagonist also known as Anakinra from Amgen, Inc.)

In a specific embodiment, compositions of the invention are administered in combination with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) or combination of one or more of the components of CHOP. In one embodiment, the compositions of the invention are administered in combination with anti-CD20 antibodies, human monoclonal anti-CD20 antibodies. In another embodiment, the compositions of the invention are administered in combination with anti-CD20 antibodies and CHOP, or anti-CD20 antibodies and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. In a specific embodiment, compositions of the invention are administered in combination with Rituximab. In a further embodiment, compositions of the invention are administered with Rituximab and CHOP, or Rituximab and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. In a specific embodiment, compositions of the invention are administered in combination with tositumomab. In a further embodiment, compositions of the invention are administered with tositumomab and CHOP, or tositumomab and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. The anti-CD20 antibodies may optionally be associated with radioisotopes, toxins or cytotoxic prodrugs.

In another specific embodiment, the compositions of the invention are administered in combination Zevalin™. In a further embodiment, compositions of the invention are administered with Zevalin™ and CHOP, or Zevalin™ and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. Zevalin™ may be associated with one or more radisotopes. Particularly preferred isotopes are ⁹⁰Y and ¹¹¹In.

In an additional embodiment, the Therapeutics of the invention are administered in combination with cytokines. Cytokines that may be administered with the Therapeutics of the invention include, but are not limited to, IL2, IL3, IL4, IL5, IL6, IL7, IL10, IL12, IL13, IL15, anti-CD40, CD40L, IFN-gamma and TNF-alpha. In another embodiment, Therapeutics of the invention may be administered with any interleukin, including, but not limited to, IL-1alpha, IL-1beta, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-19, IL-20, and IL-21.

In one embodiment, the Therapeutics of the invention are administered in combination with members of the TNF family. TNF, TNF-related or TNF-like molecules that may be administered with the Therapeutics of the invention include, but are not limited to, soluble forms of TNF-alpha, lymphotoxin-alpha (LT-alpha, also known as TNF-beta), LT-beta (found in complex heterotrimer LT-alpha2-beta), OPGL, FasL, CD27L, CD30L, CD40L, 4-1BBL, DcR3, OX40L, TNF-gamma (International Publication No. WO 96/14328), AIM-I (International Publication No. WO 97/33899), endokine-alpha (International Publication No. WO 98/07880), OPG, and neutrokine-alpha (International Publication No. WO 98/18921, OX40, and nerve growth factor (NGF), and soluble forms of Fas, CD30, CD27, CD40 and 4-IBB, TR2 (International Publication No. WO 96/34095), DR3 (International Publication No. WO 97/33904), DR4 (International Publication No. WO 98/32856), TR5 (International Publication No. WO 98/30693), TRANK, TR9 (International Publication No. WO 98/56892), TR10 (International Publication No. WO 98/54202), 312C2 (International Publication No. WO 98/06842), and TR12, and soluble forms CD154, CD70, and CD153.

In an additional embodiment, the Therapeutics of the invention are administered in combination with angiogenic proteins. Angiogenic proteins that may be administered with the Therapeutics of the invention include, but are not limited to, Glioma Derived Growth Factor (GDGF), as disclosed in European Patent Number EP-399816; Platelet Derived Growth Factor-A (PDGF-A), as disclosed in European Patent Number EP-682110; Platelet Derived Growth Factor-B (PDGF-B), as disclosed in European Patent Number EP-282317; Placental Growth Factor (PlGF), as disclosed in International Publication Number WO 92/06194; Placental Growth Factor-2 (PlGF-2), as disclosed in Hauser et al., Growth Factors, 4:259-268 (1993); Vascular Endothelial Growth Factor (VEGF), as disclosed in International Publication Number WO 90/13649; Vascular Endothelial Growth Factor-A (VEGF-A), as disclosed in European Patent Number EP-506477; Vascular Endothelial Growth Factor-2 (VEGF-2), as disclosed in International Publication Number WO 96/39515; Vascular Endothelial Growth Factor B (VEGF-3); Vascular Endothelial Growth Factor B-186 (VEGF-B186), as disclosed in International Publication Number WO 96/26736; Vascular Endothelial Growth Factor-D (VEGF-D), as disclosed in International Publication Number WO 98/02543; Vascular Endothelial Growth Factor-D (VEGF-D), as disclosed in International Publication Number WO 98/07832; and Vascular Endothelial Growth Factor-E (VEGF-E), as disclosed in German Patent Number DE19639601. The above mentioned references are herein incorporated by reference in their entireties.

In an additional embodiment, the Therapeutics of the invention are administered in combination with Fibroblast Growth Factors. Fibroblast Growth Factors that may be administered with the Therapeutics of the invention include, but are not limited to, FGF-1, FGF-2, FGF-3, FGF-4, FGF-5, FGF-6, FGF-7, FGF-8, FGF-9, FGF-10, FGF-11, FGF-12, FGF-13, FGF-14, and FGF-15.

In an additional embodiment, the Therapeutics of the invention are administered in combination with hematopoietic growth factors. Hematopoietic growth factors that may be administered with the Therapeutics of the invention include, but are not limited to, granulocyte macrophage colony stimulating factor (GM-CSF) (sargramostim, LEUKINE™, PROKINET™), granulocyte colony stimulating factor (G-CSF) (filgrastim, NEUPOGEN™), macrophage colony stimulating factor (M-CSF, CSF-1) erythropoietin (epoetin alfa, EPOGEN™, PROCRIT™), stem cell factor (SCF, c-kit ligand, steel factor), megakaryocyte colony stimulating factor, PIXY321 (a GMCSF/IL-3 fusion protein), interleukins, especially any one or more of IL-1 through IL-12, interferon-gamma, or thrombopoietin.

In certain embodiments, Therapeutics of the present invention are administered in combination with adrenergic blockers, such as, for example, acebutolol, atenolol, betaxolol, bisoprolol, carteolol, labetalol, metoprolol, nadolol, oxprenolol, penbutolol, pindolol, propranolol, sotalol, and timolol.

In another embodiment, the Therapeutics of the invention are administered in combination with an antiarrhythmic drug (e.g., adenosine, amidoarone, bretylium, digitalis, digoxin, digitoxin, diliazem, disopyramide, esmolol, flecamide, lidocaine, mexiletine, moricizine, phenyloin, procainamide, N-acetyl procainamide, propafenone, propranolol, quinidine, sotalol, tocamide, and verapamil).

In another embodiment, the Therapeutics of the invention are administered in combination with diuretic agents, such as carbonic anhydrase-inhibiting agents (e.g., acetazolamide, dichlorphenamide, and methazolamide), osmotic diuretics (e.g., glycerin, isosorbide, mannitol, and urea), diuretics that inhibit Na⁺-K⁺-2Cl⁻ symport (e.g., furosemide, bumetanide, azosemide, piretanide, tripamide, ethacrynic acid, muzolimine, and torsemide), thiazide and thiazide-like diuretics (e.g., bendroflumethiazide, benzthiazide, chlorothiazide, hydrochlorothiazide, hydroflumethiazide, methyclothiazide, polythiazide, trichormethiazide, chlorthalidone, indapamide, metolazone, and quinethazone), potassium sparing diuretics (e.g., amiloride and triamterene), and mineralcorticoid receptor antagonists (e.g., spironolactone, canrenone, and potassium canrenoate).

In one embodiment, the Therapeutics of the invention are administered in combination with treatments for endocrine and/or hormone imbalance disorders. Treatments for endocrine and/or hormone imbalance disorders include, but are not limited to, ¹²⁷I, radioactive isotopes of iodine such as ¹³¹I and ¹²³I; recombinant growth hormone, such as HUMATROPE™ (recombinant somatropin); growth hormone analogs such as PROTROPIN™ (somatrem); dopamine agonists such as PARLODEL™ (bromocriptine); somatostatin analogs such as SANDOSTATIN™ (octreotide); gonadotropin preparations such as PREGNYL™, A.P.L.™ and PROFASI™ (chorionic gonadotropin (CG)), PERGONAL™ (menotropins), and METRODIN™ (urofollitropin (uFSH)); synthetic human gonadotropin releasing hormone preparations such as FACTREL™ and LUTREPULSE™ (gonadorelin hydrochloride); synthetic gonadotropin agonists such as LUPRON™ (leuprolide acetate), SUPPRELIN™ (histrelin acetate), SYNAREL™ (nafarelin acetate), and ZOLADEX™ (goserelin acetate); synthetic preparations of thyrotropin-releasing hormone such as RELEFACT TRH™ and THYPINONE™ (protirelin); recombinant human TSH such as THYROGEN™; synthetic preparations of the sodium salts of the natural isomers of thyroid hormones such as L-T₄™, SYNTHROID™ and LEVOTHROID™ (levothyroxine sodium), L-T₃™, CYTOMEL™ and TRIOSTAT™ (liothyroine sodium), and THYROLAR™ (liotrix); antithyroid compounds such as 6-n-propylthiouracil (propylthiouracil), 1-methyl-2-mercaptoimidazole and TAPAZOLE™ (methimazole), NEO-MERCAZOLE™ (carbimazole); beta-adrenergic receptor antagonists such as propranolol and esmolol; Ca²⁺ channel blockers; dexamethasone and iodinated radiological contrast agents such as TELEPAQUE™ (iopanoic acid) and ORAGRAFIN™ (sodium ipodate).

Additional treatments for endocrine and/or hormone imbalance disorders include, but are not limited to, estrogens or congugated estrogens such as ESTRACE™ (estradiol), ESTINYL™ (ethinyl estradiol), PREMARIN™, ESTRATAB™, ORTHO-EST™, OGEN™ and estropipate (estrone), ESTROVIS™ (quinestrol), ESTRADERM™ (estradiol), DELESTROGEN™ and VALERGEN™ (estradiol valerate), DEPO-ESTRADIOL CYPIONATE™ and ESTROJECT LA™ (estradiol cypionate); antiestrogens such as NOLVADEX™ (tamoxifen), SEROPHENE™ and CLOMID™ (clomiphene); progestins such as DURALUTIN™ (hydroxyprogesterone caproate), MPA™ and DEPO-PROVERA™(medroxyprogesterone acetate), PROVERA™ and CYCRIN™ (MPA), MEGACE™ (megestrol acetate), NORLUTIN™ (norethindrone), and NORLUTATE™ and AYGESTIN™ (norethindrone acetate); progesterone implants such as NORPLANT SYSTEM™ (subdermal implants of norgestrel); antiprogestins such as RU 486™ (mifepristone); hormonal contraceptives such as ENOVID™ (norethynodrel plus mestranol), PROGESTASERT™ (intrauterine device that releases progesterone), LOESTRIN™, BREVICON™, MODICON™ GENORA™, NELONA™, NORINYL™, OVACON-35™ and OVACON-50™ (ethinyl estradiol/norethindrone), LEVLEN™, NORDETTE™, TRI-LEVLEN™ and TRIPHASIL-21™ (ethinyl estradialevonorgestrel) LO/OVRAL™ and OVRAL™ (ethinyl estradiol/norgestrel), DEIVHJLEN™ (ethinyl estradiol/ethynodiol diacetate), NORINYL™, ORTHO-NOVUM™, NORETHIN™, GENORA™, and NELOVA™ (norethindrone/mestranol), DESOGEN™ and ORTHO-CEPT™ (ethinyl estradiol/desogestrel), ORTHO-CYCLEN™ and ORTHO-TRICYCLEN™ (ethinyl estradiol/norgestimate), MICRONOR™ and NOR-QD™ (norethindrone), and OVRETTE™ (norgestrel).

Additional treatments for endocrine and/or hormone imbalance disorders include, but are not limited to, testosterone esters such as methenolone acetate and testosterone undecanoate; parenteral and oral androgens such as TESTOJECT-50™ (testosterone), TESTEX™ (testosterone propionate), DELATESTRYL™ (testosterone enanthate), DEPO-TESTOSTERONE™ (testosterone cypionate), DANOCRINE™ (danazol), HALOTESTIN™ (fluoxymesterone), ORETON METHYL™, TESTRED™ and VIRILON™ (methyltestosterone), and OXANDRIN™ (oxandrolone); testosterone transdermal systems such as TESTODERM™; androgen receptor antagonist and 5-alpha-reductase inhibitors such as ANDROCUR™ (cyproterone acetate), EULEXIN™ (flutamide), and PROSCAR™ (finasteride); adrenocorticotropic hormone preparations such as CORTROSYN™ (cosyntropin); adrenocortical steroids and their synthetic analogs such as ACLOVATE™ (alclometasone dipropionate), CYCLOCORT™ (amcinonide), BECLOVENT™ and VANCERIL™ (beclomethasone dipropionate), CELESTONE™ (betamethasone), BENISONE™ and UTICORT™ (betamethasone benzoate), DIPROSONE™ (betamethasone dipropionate), CELESTONE PHOSPHATE™ (betamethasone sodium phosphate), CELESTONE SOLUSPAN™ (betamethasone sodium phosphate and acetate), BETA-VAL™ and VALISONE™ (betamethasone valerate), TEMOVATE™ (clobetasol propionate), CLODERM™ (clocortolone pivalate), CORTEF™ and HYDROCORTONE™ (cortisol (hydrocortisone)), HYDROCORTONE ACETATE™ (cortisol (hydrocortisone) acetate), LOCOID™ (cortisol (hydrocortisone) butyrate), HYDROCORTONE PHOSPHATE™ (cortisol (hydrocortisone) sodium phosphate), A-HYDROCORT™ and SOLU CORTEF™ (cortisol (hydrocortisone) sodium succinate), WESTCORT™ (cortisol (hydrocortisone) valerate), CORTISONE ACETATE™ (cortisone acetate), DESOWEN™ and TRIDESILON™ (desonide), TOPICORT™ (desoximetasone), DECADRON™ (dexamethasone), DECADRON LA™ (dexamethasone acetate), DECADRON PHOSPHATE™ and HEXADROL PHOSPHATE™ (dexamethasone sodium phosphate), FLORONE™ and MAXIFLOR™ (diflorasone diacetate), FLORINEF ACETATE™ (fludrocortisone acetate), AEROBID™ and NASALIDE™ (flunisolide), FLUONID™ and SYNALAR™ (fluocinolone acetonide), LIDEX™ (fluocinonide), FLUOR-OP™ and FML™ (fluorometholone), CORDRAN™ (flurandrenolide), HALOG™ (halcinonide), HMS LIZUIFILM™ (medrysone), MEDROL™ (methylprednisolone), DEPO-MEDROL™ and MEDROL ACETATE™ (methylprednisone acetate), A-METHAPRED™ and SOLUMEDROL™ (methylprednisolone sodium succinate), ELOCON™ (mometasone furoate), HALDRONE™ (paramethasone acetate), DELTA-CORTEF™ (prednisolone), ECONOPRED™ (prednisolone acetate), HYDELTRASOL™ (prednisolone sodium phosphate), HYDELTRA-T.B.A™ (prednisolone tebutate), DELTASONE™ (prednisone), ARISTOCORT™ and KENACORT™ (triamcinolone), KENALOG™ (triamcinolone acetonide), ARISTOCORT™ and KENACORT DIACETATE™ (triamcinolone diacetate), and ARISTOSPAN™ (triamcinolone hexacetonide); inhibitors of biosynthesis and action of adrenocortical steroids such as CYTADREN™ (aminoglutethimide), NIZORAL™ (ketoconazole), MODRASTANE™ (trilostane), and METOPIRONE™ (metyrapone); bovine, porcine or human insulin or mixtures thereof; insulin analogs; recombinant human insulin such as HUMULIN™ and NOVOLIN™; oral hypoglycemic agents such as ORAMIDE™ and ORINASE™ (tolbutamide), DIABINESE™ (chlorpropamide), TOLAMIDE™ and TOLINASE™ (tolazamide), DYMELOR™ (acetohexamide), glibenclamide, MICRONASE™, DIBETA™ and GLYNASE™ (glyburide), GLUCOTROL™ (glipizide), and DIAMICRON™ (gliclazide), GLUCOPHAGE™ (metformin), ciglitazone, pioglitazone, and alpha-glucosidase inhibitors; bovine or porcine glucagon; somatostatins such as SANDOSTATIN™ (octreotide); and diazoxides such as PROGLYCEM™ (diazoxide).

In one embodiment, the Therapeutics of the invention are administered in combination with treatments for uterine motility disorders. Treatments for uterine motility disorders include, but are not limited to, estrogen drugs such as conjugated estrogens (e.g., PREMARIN® and ESTRATAB®), estradiols (e.g., CLIMARA® and ALORA®), estropipate, and chlorotrianisene; progestin drugs (e.g., AMEN® (medroxyprogesterone), MICRONOR® (norethidrone acetate), PROMETRIUM® progesterone, and megestrol acetate); and estrogen/progesterone combination therapies such as, for example, conjugated estrogens/medroxyprogesterone (e.g., PREMPRO™ and PREMPHASE®) and norethindrone acetate/ethinyl estsradiol (e.g., FEMHRTT™).

In an additional embodiment, the Therapeutics of the invention are administered in combination with drugs effective in treating iron deficiency and hypochromic anemias, including but not limited to, ferrous sulfate (iron sulfate, FEOSOL™), ferrous fumarate (e.g., FEOSTAT™), ferrous gluconate (e.g., FERGON™), polysaccharide-iron complex (e.g., NIFEREX™), iron dextran injection (e.g., INFED™), cupric sulfate, pyroxidine, riboflavin, Vitamin B₁₂, cyancobalamin injection (e.g., REDISOL™, RUBRAMIN PC™) hydroxocobalamin, folic acid (e.g., FOLVITE™), leucovorin (folinic acid, 5-CHOH4PteGlu, citrovorum factor) or WELLCOVORIN (Calcium salt of leucovorin), transferrin or ferritin.

In other embodiments, the Therapeutics of the invention are administered in combination with agents used to treat neurological disorders. Neurological agents that may be administered with the Therapeutics of the invention include, but are not limited to, antiepileptic agents (e.g., carbamazepine, clonazepam, ethosuximide, phenobarbital, phenyloin, primidone, valproic acid, divalproex sodium, felbamate, gabapentin, lamotrigine, levetiracetam, oxcarbazepine, tiagabine, topiramate, zonisamide, diazepam, lorazepam, and clonazepam), antiparkinsonian agents (e.g., levodopa/carbidopa, selegiline, amantidine, bromocriptine, pergolide, ropinirole, pramipexole, benztropine; biperiden; ethopropazine; procyclidine; trihexyphenidyl, tolcapone), and ALS therapeutics (e.g. riluzole).

In another embodiment, Therapeutics of the invention are administered in combination with vasodilating agents and/or calcium channel blocking agents. Vasodilating agents that may be administered with the Therapeutics of the invention include, but are not limited to, Angiotensin Converting Enzyme (ACE) inhibitors (e.g., papaverine, isoxsuprine, benazepril, captopril, cilazapril, enalapril, enalaprilat, fosinopril, lisinopril, moexipril, perindopril, quinapril, ramipril, spirapril, trandolapril, and nylidrin), and nitrates (e.g., isosorbide dinitrate, isosorbide mononitrate, and nitroglycerin). Examples of calcium channel blocking agents that may be administered in combination with the Therapeutics of the invention include, but are not limited to amlodipine, bepridil, diltiazem, felodipine, flunarizine, isradipine, nicardipine, nifedipine, nimodipine, and verapamil.

In certain embodiments, the Therapeutics of the invention are administered in combination with treatments for gastrointestinal disorders. Treatments for gastrointestinal disorders that may be administered with the Therapeutic of the invention include, but are not limited to, H₂ histamine receptor antagonists (e.g., TAGAMET™ (cimetidine), ZANTAC™ (ranitidine), PEPCID™ (famotidine), and AXID™ (nizatidine)); inhibitors of ATPase (e.g., PREVACID™ (lansoprazole) and PRILOSEC™ (omeprazole)); Bismuth compounds (e.g., PEPTO-BISMOL™ (bismuth subsalicylate) and DE-NOL™ (bismuth subcitrate)); various antacids; sucralfate; prostaglandin analogs (e.g. CYTOTEC™ (misoprostol)); muscarinic cholinergic antagonists; laxatives (e.g., surfactant laxatives, stimulant laxatives, saline and osmotic laxatives); antidiarrheal agents (e.g., LOMOTIL™ (diphenoxylate), MOTOFEN™ (diphenoxin), and IMODIUM™ (loperamide hydrochloride)), synthetic analogs of somatostatin such as SANDOSTATIN™ (octreotide), antiemetic agents (e.g., ZOFRAN™ (ondansetron), KYTRIL™ (granisetron hydrochloride), tropisetron, dolasetron, metoclopramide, chlorpromazine, perphenazine, prochlorperazine, promethazine, thiethylperazine, triflupromazine, domperidone, haloperidol, droperidol, trimethobenzamide, dexamethasone, methylprednisolone, dronabinol, and nabilone); D2 antagonists (e.g., metoclopramide, trimethobenzamide and chlorpromazine); bile salts; chenodeoxycholic acid; ursodeoxycholic acid; and pancreatic enzyme preparations such as pancreatin and pancrelipase.

In additional embodiments, the Therapeutics of the invention are administered in combination with other therapeutic or prophylactic regimens, such as, for example, radiation therapy.

Example 14 Method of Treating Decreased Levels of the Polypeptide

The present invention relates to a method for treating an individual in need of an increased level of a polypeptide of the invention in the body comprising administering to such an individual a composition comprising a therapeutically effective amount of polypeptides (including agonists thereto), and/or antibodies of the invention. Moreover, it will be appreciated that conditions caused by a decrease in the standard or normal expression level of a polypeptide of the present invention in an individual may be treated by administering agonists of said polypeptide. Thus, the invention also provides a method of treatment of an individual in need of an increased level of the polypeptide comprising administering to such an individual a Therapeutic comprising an amount of the agonist (including polypeptides and antibodies of the present invention) to increase the activity level of the polypeptide in such an individual.

For example, a patient with decreased levels of a polypeptide receives a daily dose 0.1-100 μg/kg of the agonist for six consecutive days. The exact details of the dosing scheme, based on administration and formulation, are provided in Example 13.

Example 15 Method of Treating Increased Levels of the Polypeptide

The present invention also relates to a method of treating an individual in need of a decreased level of a polypeptide of the invention in the body comprising administering to such an individual a composition comprising a therapeutically effective amount of an antagonist of the invention (including polypeptides and antibodies of the invention).

In one example, antisense technology is used to inhibit production of a polypeptide of the present invention. This technology is one example of a method of decreasing levels of a polypeptide, due to a variety of etiologies, such as cancer.

For example, a patient diagnosed with abnormally increased levels of a polypeptide is administered intravenously antisense polynucleotides at 0.5, 1.0, 1.5, 2.0 and 3.0 mg/kg day for 21 days. This treatment is repeated after a 7-day rest period if the treatment was well tolerated. The antisense polynucleotides of the present invention can be formulated using techniques and formulations described herein (e.g. see Example 13), or otherwise known in the art.

Example 16 Method of Treatment Using Gene Therapy-Ex Vivo

One method of gene therapy transplants fibroblasts, which are capable of expressing a polypeptide, onto a patient. Generally, fibroblasts are obtained from a subject by skin biopsy. The resulting tissue is placed in tissue-culture medium and separated into small pieces. Small chunks of the tissue are placed on a wet surface of a tissue culture flask, approximately ten pieces are placed in each flask. The flask is turned upside down, closed tight and left at room temperature over night. After 24 hours at room temperature, the flask is inverted and the chunks of tissue remain fixed to the bottom of the flask and fresh media (e.g., Ham's F12 media, with 10% FBS, penicillin and streptomycin) is added. The flasks are then incubated at 37 degree C. for approximately one week.

At this time, fresh media is added and subsequently changed every several days. After an additional two weeks in culture, a monolayer of fibroblasts emerge. The monolayer is trypsinized and scaled into larger flasks.

pMV-7 (Kirschmeier, P. T. et al., DNA, 7:219-25 (1988)), flanked by the long terminal repeats of the Moloney murine sarcoma virus, is digested with EcoRI and HindIII and subsequently treated with calf intestinal phosphatase. The linear vector is fractionated on agarose gel and purified, using glass beads.

The cDNA encoding a polypeptide of the present invention can be amplified using PCR primers which correspond to the 5′ and 3′ end sequences respectively as set forth in Example 1 using primers and having appropriate restriction sites and initiation/stop codons, if necessary. Preferably, the 5′ primer contains an EcoRI site and the 3′ primer includes a HindIII site. Equal quantities of the Moloney murine sarcoma virus linear backbone and the amplified EcoRI and HindIII fragment are added together, in the presence of T4 DNA ligase. The resulting mixture is maintained under conditions appropriate for ligation of the two fragments. The ligation mixture is then used to transform bacteria HB101, which are then plated onto agar containing kanamycin for the purpose of confirming that the vector has the gene of interest properly inserted.

The amphotropic pA317 or GP+am12 packaging cells are grown in tissue culture to confluent density in Dulbecco's Modified Eagles Medium (DMEM) with 10% calf serum (CS), penicillin and streptomycin. The MSV vector containing the gene is then added to the media and the packaging cells transduced with the vector. The packaging cells now produce infectious viral particles containing the gene (the packaging cells are now referred to as producer cells).

Fresh media is added to the transduced producer cells, and subsequently, the media is harvested from a 10 cm plate of confluent producer cells. The spent media, containing the infectious viral particles, is filtered through a millipore filter to remove detached producer cells and this media is then used to infect fibroblast cells. Media is removed from a sub-confluent plate of fibroblasts and quickly replaced with the media from the producer cells. This media is removed and replaced with fresh media. If the titer of virus is high, then virtually all fibroblasts will be infected and no selection is required. If the titer is very low, then it is necessary to use a retroviral vector that has a selectable marker, such as neo or his. Once the fibroblasts have been efficiently infected, the fibroblasts are analyzed to determine whether protein is produced.

The engineered fibroblasts are then transplanted onto the host, either alone or after having been grown to confluence on cytodex 3 microcarrier beads.

Example 17 Gene Therapy Using Endogenous Genes Corresponding to Polynucleotides of the Invention

Another method of gene therapy according to the present invention involves operably associating the endogenous polynucleotide sequence of the invention with a promoter via homologous recombination as described, for example, in U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication NO: WO 96/29411, published Sep. 26, 1996; International Publication NO: WO 94/12650, published Aug. 4, 1994; Koller et al., Proc. Natl. Acad. Sci. USA, 86:8932-8935 (1989); and Zijlstra et al., Nature, 342:435-438 (1989). This method involves the activation of a gene which is present in the target cells, but which is not expressed in the cells, or is expressed at a lower level than desired.

Polynucleotide constructs are made which contain a promoter and targeting sequences, which are homologous to the 5′ non-coding sequence of endogenous polynucleotide sequence, flanking the promoter. The targeting sequence will be sufficiently near the 5′ end of the polynucleotide sequence so the promoter will be operably linked to the endogenous sequence upon homologous recombination. The promoter and the targeting sequences can be amplified using PCR. Preferably, the amplified promoter contains distinct restriction enzyme sites on the 5′ and 3′ ends. Preferably, the 3′ end of the first targeting sequence contains the same restriction enzyme site as the 5′ end of the amplified promoter and the 5′ end of the second targeting sequence contains the same restriction site as the 3′ end of the amplified promoter.

The amplified promoter and the amplified targeting sequences are digested with the appropriate restriction enzymes and subsequently treated with calf intestinal phosphatase. The digested promoter and digested targeting sequences are added together in the presence of T4 DNA ligase. The resulting mixture is maintained under conditions appropriate for ligation of the two fragments. The construct is size fractionated on an agarose gel, then purified by phenol extraction and ethanol precipitation.

In this Example, the polynucleotide constructs are administered as naked polynucleotides via electroporation. However, the polynucleotide constructs may also be administered with transfection-facilitating agents, such as liposomes, viral sequences, viral particles, precipitating agents, etc. Such methods of delivery are known in the art.

Once the cells are transfected, homologous recombination will take place which results in the promoter being operably linked to the endogenous polynucleotide sequence. This results in the expression of polynucleotide corresponding to the polynucleotide in the cell. Expression may be detected by immunological staining, or any other method known in the art.

Fibroblasts are obtained from a subject by skin biopsy. The resulting tissue is placed in DMEM+10% fetal calf serum. Exponentially growing or early stationary phase fibroblasts are trypsinized and rinsed from the plastic surface with nutrient medium. An aliquot of the cell suspension is removed for counting, and the remaining cells are subjected to centrifugation. The supernatant is aspirated and the pellet is resuspended in 5 ml of electroporation buffer (20 mM HEPES pH 7.3, 137 mM NaCl, 5 mM KCl, 0.7 mM Na_(z) HPO₄, 6 mM dextrose). The cells are recentrifuged, the supernatant aspirated, and the cells resuspended in electroporation buffer containing 1 mg/ml acetylated bovine serum albumin. The final cell suspension contains approximately 3×10⁶ cells/ml. Electroporation should be performed immediately following resuspension.

Plasmid DNA is prepared according to standard techniques. For example, to construct a plasmid for targeting to the locus corresponding to the polynucleotide of the invention, plasmid pUC18 (MBI Fermentas, Amherst, N.Y.) is digested with HindIII. The CMV promoter is amplified by PCR with an XbaI site on the 5′ end and a BamHI site on the 3′ end. Two non-coding sequences are amplified via PCR: one non-coding sequence (fragment 1) is amplified with a HindIII site at the 5′ end and an Xba site at the 3′ end; the other non-coding sequence (fragment 2) is amplified with a BamHI site at the 5′ end and a HindIII site at the 3′ end. The CMV promoter and the fragments (1 and 2) are digested with the appropriate enzymes (CMV promoter—XbaI and BamHI; fragment 1-XbaI; fragment 2-BamHI) and ligated together. The resulting ligation product is digested with HindIII, and ligated with the HindIII-digested pUC18 plasmid.

Plasmid DNA is added to a sterile cuvette with a 0.4 cm electrode gap (Bio-Rad). The final DNA concentration is generally at least 120 μg/ml. 0.5 ml of the cell suspension (containing approximately 1.5×10⁶ cells) is then added to the cuvette, and the cell suspension and DNA solutions are gently mixed. Electroporation is performed with a Gene-Pulser apparatus (Bio-Rad). Capacitance and voltage are set at 960 μF and 250-300 V, respectively. As voltage increases, cell survival decreases, but the percentage of surviving cells that stably incorporate the introduced DNA into their genome increases dramatically. Given these parameters, a pulse time of approximately 14-20 mSec should be observed.

Electroporated cells are maintained at room temperature for approximately 5 min, and the contents of the cuvette are then gently removed with a sterile transfer pipette. The cells are added directly to 10 ml of prewarmed nutrient media (DMEM with 15% calf serum) in a 10 cm dish and incubated at 37 degree C. The following day, the media is aspirated and replaced with 10 ml of fresh media and incubated for a further 16-24 hours.

The engineered fibroblasts are then injected into the host, either alone or after having been grown to confluence on cytodex 3 microcarrier beads. The fibroblasts now produce the protein product. The fibroblasts can then be introduced into a patient as described above.

Example 18 Method of Treatment Using Gene Therapy—In Vivo

Another aspect of the present invention is using in vivo gene therapy methods to prevent, treat, and/or ameliorate immune diseases and disorders. The gene therapy method relates to the introduction of naked nucleic acid (DNA, RNA, and antisense DNA or RNA) sequences into an animal to increase or decrease the expression of the polypeptide. The polynucleotide of the present invention may be operatively linked to (i.e., associated with) a promoter or any other genetic elements necessary for the expression of the polypeptide by the target tissue. Such gene therapy and delivery techniques and methods are known in the art, see, for example, WO90/11092, WO98/11779; U.S. Pat. Nos. 5,693,622, 5,705,151, 5,580,859; Tabata et al., Cardiovasc. Res. 35(3):470-479 (1997); Chao et al., Pharmacol. Res. 35(6):517-522 (1997); Wolff, Neuromuscul. Disord. 7(5):314-318 (1997); Schwartz et al., Gene Ther. 3 (5): 405-411 (1996); Tsurumi et al., Circulation 94 (12):3281-3290 (1996) (incorporated herein by reference).

The polynucleotide constructs may be delivered by any method that delivers injectable materials to the cells of an animal, such as, injection into the interstitial space of tissues (heart, muscle, skin, lung, liver, intestine and the like). The polynucleotide constructs can be delivered in a pharmaceutically acceptable liquid or aqueous carrier.

The term “naked” polynucleotide, DNA or RNA, refers to sequences that are free from any delivery vehicle that acts to assist, promote, or facilitate entry into the cell, including viral sequences, viral particles, liposome formulations, LIPOFECTIN™ or precipitating agents and the like. However, the polynucleotides of the present invention may also be delivered in liposome formulations (such as those taught in Felgner P. L. et al. (1995) Ann. NY Acad. Sci. 772:126-139 and Abdallah B. et al. (1995) Biol. Cell 85(1):1-7) which can be prepared by methods well known to those skilled in the art.

The polynucleotide vector constructs used in the gene therapy method are preferably constructs that will not integrate into the host genome nor will they contain sequences that allow for replication. Any strong promoter known to those skilled in the art can be used for driving the expression of DNA. Unlike other gene therapy techniques, one major advantage of introducing naked nucleic acid sequences into target cells is the transitory nature of the polynucleotide synthesis in the cells. Studies have shown that non-replicating DNA sequences can be introduced into cells to provide production of the desired polypeptide for periods of up to six months.

The polynucleotide construct can be delivered to the interstitial space of tissues within an animal, including muscle, skin, brain, lung, liver, spleen, bone marrow, thymus, heart, lymph, blood, bone, cartilage, pancreas, kidney, gall bladder, stomach, intestine, testis, ovary, uterus, rectum, nervous system, eye, gland, and connective tissue. Interstitial space of the tissues comprises the intercellular fluid, mucopolysaccharide matrix among the reticular fibers of organ tissues, elastic fibers in the walls of vessels or chambers, collagen fibers of fibrous tissues, or that same matrix within connective tissue ensheathing muscle cells or in the lacunae of bone. It is similarly the space occupied by the plasma of the circulation and the lymph fluid of the lymphatic channels. Delivery to the interstitial space of muscle tissue is preferred for the reasons discussed below. They may be conveniently delivered by injection into the tissues comprising these cells. They are preferably delivered to and expressed in persistent, non-dividing cells which are differentiated, although delivery and expression may be achieved in non-differentiated or less completely differentiated cells, such as, for example, stem cells of blood or skin fibroblasts. In vivo muscle cells are particularly competent in their ability to take up and express polynucleotides.

For the naked polynucleotide injection, an effective dosage amount of DNA or RNA will be in the range of from about 0.05 g/kg body weight to about 50 mg/kg body weight. Preferably the dosage will be from about 0.005 mg/kg to about 20 mg/kg and more preferably from about 0.05 mg/kg to about 5 mg/kg. Of course, as the artisan of ordinary skill will appreciate, this dosage will vary according to the tissue site of injection. The appropriate and effective dosage of nucleic acid sequence can readily be determined by those of ordinary skill in the art and may depend on the condition being treated and the route of administration. The preferred route of administration is by the parenteral route of injection into the interstitial space of tissues. However, other parenteral routes may also be used, such as, inhalation of an aerosol formulation particularly for delivery to lungs or bronchial tissues, throat or mucous membranes of the nose. In addition, naked polynucleotide constructs can be delivered to arteries during angioplasty by the catheter used in the procedure.

The dose response effects of injected polynucleotide in muscle in vivo is determined as follows. Suitable template DNA for production of mRNA coding for polypeptide of the present invention is prepared in accordance with a standard recombinant DNA methodology. The template DNA, which may be either circular or linear, is either used as naked DNA or complexed with liposomes. The quadriceps muscles of mice are then injected with various amounts of the template DNA.

Five to six week old female and male Balb/C mice are anesthetized by intraperitoneal injection with 0.3 ml of 2.5% Avertin. A 1.5 cm incision is made on the anterior thigh, and the quadriceps muscle is directly visualized. The template DNA is injected in 0.1 ml of carrier in a 1 cc syringe through a 27 gauge needle over one minute, approximately 0.5 cm from the distal insertion site of the muscle into the knee and about 0.2 cm deep. A suture is placed over the injection site for future localization, and the skin is closed with stainless steel clips.

After an appropriate incubation time (e.g., 7 days) muscle extracts are prepared by excising the entire quadriceps. Every fifth 15 μm cross-section of the individual quadriceps muscles is histochemically stained for protein expression. A time course for protein expression may be done in a similar fashion except that quadriceps from different mice are harvested at different times. Persistence of DNA in muscle following injection may be determined by Southern blot analysis after preparing total cellular DNA and HIRT supernatants from injected and control mice. The results of the above experimentation in mice can be used to extrapolate proper dosages and other treatment parameters in humans and other animals using naked DNA.

Example 19 Transgenic Animals

The polypeptides of the invention can also be expressed in transgenic animals. Animals of any species, including, but not limited to, mice, rats, rabbits, hamsters, guinea pigs, pigs, micro-pigs, goats, sheep, cows and non-human primates, e.g., baboons, monkeys, and chimpanzees may be used to generate transgenic animals. In a specific embodiment, techniques described herein or otherwise known in the art, are used to express polypeptides of the invention in humans, as part of a gene therapy protocol.

Any technique known in the art may be used to introduce the transgene (i.e., polynucleotides of the invention) into animals to produce the founder lines of transgenic animals. Such techniques include, but are not limited to, pronuclear microinjection (Paterson et al., Appl. Microbiol. Biotechnol. 40:691-698 (1994); Carver et al., Biotechnology (NY) 11:1263-1270 (1993); Wright et al., Biotechnology (NY) 9:830-834 (1991); and Hoppe et al., U.S. Pat. No. 4,873,191 (1989)); retrovirus mediated gene transfer into germ lines (Van der Putten et al., Proc. Natl. Acad. Sci., USA 82:6148-6152 (1985)), blastocysts or embryos; gene targeting in embryonic stem cells (Thompson et al., Cell 56:313-321 (1989)); electroporation of cells or embryos (Lo, 1983, Mol. Cell. Biol. 3:1803-1814 (1983)); introduction of the polynucleotides of the invention using a gene gun (see, e.g., Ulmer et al., Science 259:1745 (1993); introducing nucleic acid constructs into embryonic pleuripotent stem cells and transferring the stem cells back into the blastocyst; and sperm-mediated gene transfer (Lavitrano et al., Cell 57:717-723 (1989); etc. For a review of such techniques, see Gordon, “Transgenic Animals,” Intl. Rev. Cytol. 115:171-229 (1989), which is incorporated by reference herein in its entirety.

Any technique known in the art may be used to produce transgenic clones containing polynucleotides of the invention, for example, nuclear transfer into enucleated oocytes of nuclei from cultured embryonic, fetal, or adult cells induced to quiescence (Campell et al., Nature 380:64-66 (1996); Wilmut et al., Nature 385:810-813 (1997)).

The present invention provides for transgenic animals that carry the transgene in all their cells, as well as animals which carry the transgene in some, but not all their cells, i.e., mosaic animals or chimeric. The transgene may be integrated as a single transgene or as multiple copies such as in concatamers, e.g., head-to-head tandems or head-to-tail tandems. The transgene may also be selectively introduced into and activated in a particular cell type by following, for example, the teaching of Lasko et al. (Lasko et al., Proc. Natl. Acad. Sci. USA 89:6232-6236 (1992)). The regulatory sequences required for such a cell-type specific activation will depend upon the particular cell type of interest, and will be apparent to those of skill in the art. When it is desired that the polynucleotide transgene be integrated into the chromosomal site of the endogenous gene, gene targeting is preferred. Briefly, when such a technique is to be utilized, vectors containing some nucleotide sequences homologous to the endogenous gene are designed for the purpose of integrating, via homologous recombination with chromosomal sequences, into and disrupting the function of the nucleotide sequence of the endogenous gene. The transgene may also be selectively introduced into a particular cell type, thus inactivating the endogenous gene in only that cell type, by following, for example, the teaching of Gu et al. (Gu et al., Science 265:103-106 (1994)). The regulatory sequences required for such a cell-type specific inactivation will depend upon the particular cell type of interest, and will be apparent to those of skill in the art.

Once transgenic animals have been generated, the expression of the recombinant gene may be assayed utilizing standard techniques. Initial screening may be accomplished by Southern blot analysis or PCR techniques to analyze animal tissues to verify that integration of the transgene has taken place. The level of mRNA expression of the transgene in the tissues of the transgenic animals may also be assessed using techniques which include, but are not limited to, Northern blot analysis of tissue samples obtained from the animal, in situ hybridization analysis, and reverse transcriptase-PCR (rt-PCR). Samples of transgenic gene-expressing tissue may also be evaluated immunocytochemically or immunohistochemically using antibodies specific for the transgene product.

Once the founder animals are produced, they may be bred, inbred, outbred, or crossbred to produce colonies of the particular animal. Examples of such breeding strategies include, but are not limited to: outbreeding of founder animals with more than one integration site in order to establish separate lines; inbreeding of separate lines in order to produce compound transgenics that express the transgene at higher levels because of the effects of additive expression of each transgene; crossing of heterozygous transgenic animals to produce animals homozygous for a given integration site in order to both augment expression and eliminate the need for screening of animals by DNA analysis; crossing of separate homozygous lines to produce compound heterozygous or homozygous lines; and breeding to place the transgene on a distinct background that is appropriate for an experimental model of interest.

Transgenic animals of the invention have uses which include, but are not limited to, animal model systems useful in elaborating the biological function of polypeptides of the present invention, studying conditions and/or disorders associated with aberrant expression, and in screening for compounds effective in ameliorating such conditions and/or disorders.

Example 20 Knock-Out Animals

Endogenous gene expression can also be reduced by inactivating or “knocking out” the gene and/or its promoter using targeted homologous recombination. (e.g., see Smithies et al., Nature 317:230-234 (1985); Thomas & Capecchi, Cell 51:503-512 (1987); Thompson et al., Cell 5:313-321 (1989); each of which is incorporated by reference herein in its entirety). For example, a mutant, non-functional polynucleotide of the invention (or a completely unrelated DNA sequence) flanked by DNA homologous to the endogenous polynucleotide sequence (either the coding regions or regulatory regions of the gene) can be used, with or without a selectable marker and/or a negative selectable marker, to transfect cells that express polypeptides of the invention in vivo. In another embodiment, techniques known in the art are used to generate knockouts in cells that contain, but do not express the gene of interest. Insertion of the DNA construct, via targeted homologous recombination, results in inactivation of the targeted gene. Such approaches are particularly suited in research and agricultural fields where modifications to embryonic stem cells can be used to generate animal offspring with an inactive targeted gene (e.g., see Thomas & Capecchi 1987 and Thompson 1989, supra). However this approach can be routinely adapted for use in humans provided the recombinant DNA constructs are directly administered or targeted to the required site in vivo using appropriate viral vectors that will be apparent to those of skill in the art.

In further embodiments of the invention, cells that are genetically engineered to express the polypeptides of the invention, or alternatively, that are genetically engineered not to express the polypeptides of the invention (e.g., knockouts) are administered to a patient in vivo. Such cells may be obtained from the patient (i.e., animal, including human) or an MHC compatible donor and can include, but are not limited to fibroblasts, bone marrow cells, blood cells (e.g., lymphocytes), adipocytes, muscle cells, endothelial cells etc. The cells are genetically engineered in vitro using recombinant DNA techniques to introduce the coding sequence of polypeptides of the invention into the cells, or alternatively, to disrupt the coding sequence and/or endogenous regulatory sequence associated with the polypeptides of the invention, e.g., by transduction (using viral vectors, and preferably vectors that integrate the transgene into the cell genome) or transfection procedures, including, but not limited to, the use of plasmids, cosmids, YACs, naked DNA, electroporation, liposomes, etc. The coding sequence of the polypeptides of the invention can be placed under the control of a strong constitutive or inducible promoter or promoter/enhancer to achieve expression, and preferably secretion, of the polypeptides of the invention. The engineered cells which express and preferably secrete the polypeptides of the invention can be introduced into the patient systemically, e.g., in the circulation, or intraperitoneally.

Alternatively, the cells can be incorporated into a matrix and implanted in the body, e.g., genetically engineered fibroblasts can be implanted as part of a skin graft; genetically engineered endothelial cells can be implanted as part of a lymphatic or vascular graft. (See, for example, Anderson et al. U.S. Pat. No. 5,399,349; and Mulligan & Wilson, U.S. Pat. No. 5,460,959 each of which is incorporated by reference herein in its entirety).

When the cells to be administered are non-autologous or non-MHC compatible cells, they can be administered using well known techniques which prevent the development of a host immune response against the introduced cells. For example, the cells may be introduced in an encapsulated form that, while allowing for an exchange of components with the immediate extracellular environment, does not allow the introduced cells to be recognized by the host immune system.

Transgenic and “knock-out” animals of the invention have uses which include, but are not limited to, animal model systems useful in elaborating the biological function of polypeptides of the present invention, studying conditions and/or disorders associated with aberrant expression, and in screening for compounds effective in ameliorating such conditions and/or disorders.

Example 21 Assays Detecting Stimulation or Inhibition of B Cell Proliferation and Differentiation

Generation of functional humoral immune responses requires both soluble and cognate signaling between B-lineage cells and their microenvironment. Signals may impart a positive stimulus that allows a B-lineage cell to continue its programmed development, or a negative stimulus that instructs the cell to arrest its current developmental pathway. To date, numerous stimulatory and inhibitory signals have been found to influence B cell responsiveness including IL-2, IL-4, IL-5, IL-6, IL-7, IL10, IL-13, IL-14 and IL-15. Interestingly, these signals are by themselves weak effectors but can, in combination with various co-stimulatory proteins, induce activation, proliferation, differentiation, homing, tolerance and death among B cell populations.

One of the best studied classes of B-cell co-stimulatory proteins is the TNF-superfamily. Within this family CD40, CD27, and CD30 along with their respective ligands CD154, CD70, and CD153 have been found to regulate a variety of immune responses. Assays which allow for the detection and/or observation of the proliferation and differentiation of these B-cell populations and their precursors are valuable tools in determining the effects various proteins may have on these B-cell populations in terms of proliferation and differentiation. Listed below are two assays designed to allow for the detection of the differentiation, proliferation, or inhibition of B-cell populations and their precursors.

In Vitro Assay—Agonists or antagonists of the invention can be assessed for its ability to induce activation, proliferation, differentiation or inhibition and/or death in B-cell populations and their precursors. The activity of the agonists or antagonists of the invention on purified human tonsillar B cells, measured qualitatively over the dose range from 0.1 to 10,000 ng/mL, is assessed in a standard B-lymphocyte co-stimulation assay in which purified tonsillar B cells are cultured in the presence of either formalin-fixed Staphylococcus aureus Cowan I (SAC) or immobilized anti-human IgM antibody as the priming agent. Second signals such as IL-2 and IL-15 synergize with SAC and IgM crosslinking to elicit B cell proliferation as measured by tritiated-thymidine incorporation. Novel synergizing agents can be readily identified using this assay. The assay involves isolating human tonsillar B cells by magnetic bead (MACS) depletion of CD3-positive cells. The resulting cell population is greater than 95% B cells as assessed by expression of CD45R(B220).

Various dilutions of each sample are placed into individual wells of a 96-well plate to which are added 10⁵ B-cells suspended in culture medium (RPMI 1640 containing 10% FBS, 5×10⁻⁵M 2ME, 100 U/ml penicillin, 10 ug/ml streptomycin, and 10⁻⁵ dilution of SAC) in a total volume of 150 ul. Proliferation or inhibition is quantitated by a 20 h pulse (1 uCi/well) with 3H-thymidine (6.7 Ci/mM) beginning 72 h post factor addition. The positive and negative controls are IL2 and medium respectively.

In vivo Assay-BALB/c mice are injected (i.p.) twice per day with buffer only, or 2 mg/Kg of agonists or antagonists of the invention, or truncated forms thereof. Mice receive this treatment for 4 consecutive days, at which time they are sacrificed and various tissues and serum collected for analyses. Comparison of H&E sections from normal spleens and spleens treated with agonists or antagonists of the invention identify the results of the activity of the agonists or antagonists on spleen cells, such as the diffusion of peri-arterial lymphatic sheaths, and/or significant increases in the nucleated cellularity of the red pulp regions, which may indicate the activation of the differentiation and proliferation of B-cell populations Immunohistochemical studies using a B cell marker, anti-CD45R(B220), are used to determine whether any physiological changes to splenic cells, such as splenic disorganization, are due to increased B-cell representation within loosely defined B-cell zones that infiltrate established T-cell regions.

Flow cytometric analyses of the spleens from mice treated with agonist or antagonist is used to indicate whether the agonists or antagonists specifically increases the proportion of ThB+, CD45R(B220) dull B cells over that which is observed in control mice.

Likewise, a predicted consequence of increased mature B-cell representation in vivo is a relative increase in serum Ig titers. Accordingly, serum IgM and IgA levels are compared between buffer and agonists or antagonists-treated mice.

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 22 T Cell Proliferation Assay

A CD3-induced proliferation assay is performed on PBMCs and is measured by the uptake of ³H-thymidine. The assay is performed as follows. Ninety-six well plates are coated with 100 μl/well of mAb to CD3 (HIT3a, Pharmingen) or isotype-matched control mAb (B33.1) overnight at 4 degrees C. (1 μg/ml in 0.05M bicarbonate buffer, pH 9.5), then washed three times with PBS. PBMC are isolated by F/H gradient centrifugation from human peripheral blood and added to quadruplicate wells (5×10⁴/well) of mAb coated plates in RPMI containing 10% FCS and P/S in the presence of varying concentrations of agonists or antagonists of the invention (total volume 200 μl). Relevant protein buffer and medium alone are controls. After 48 hr. culture at 37 degrees C., plates are spun for 2 min. at 1000 rpm and 100 μl of supernatant is removed and stored −20 degrees C. for measurement of IL-2 (or other cytokines) if effect on proliferation is observed. Wells are supplemented with 100 μl of medium containing 0.5 μCi of ³H-thymidine and cultured at 37 degrees C. for 18-24 hr. Wells are harvested and incorporation of ³H-thymidine used as a measure of proliferation. Anti-CD3 alone is the positive control for proliferation. IL-2 (100 U/ml) is also used as a control which enhances proliferation. Control antibody which does not induce proliferation of T cells is used as the negative control for the effects of agonists or antagonists of the invention.

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 23 Effect of Agonists or Antagonists of the Invention on the Expression of MHC Class II, Costimulatory and Adhesion Molecules and Cell Differentiation of Monocytes and Monocyte-Derived Human Dendritic Cells

Dendritic cells are generated by the expansion of proliferating precursors found in the peripheral blood: adherent PBMC or elutriated monocytic fractions are cultured for 7-10 days with GM-CSF (50 ng/ml) and IL-4 (20 ng/ml). These dendritic cells have the characteristic phenotype of immature cells (expression of CD1, CD80, CD86, CD40 and MHC class II antigens). Treatment with activating factors, such as TNF-α, causes a rapid change in surface phenotype (increased expression of MHC class I and II, costimulatory and adhesion molecules, downregulation of FCγRII, upregulation of CD83). These changes correlate with increased antigen-presenting capacity and with functional maturation of the dendritic cells.

FACS analysis of surface antigens is performed as follows. Cells are treated 1-3 days with increasing concentrations of agonist or antagonist of the invention or LPS (positive control), washed with PBS containing 1% BSA and 0.02 mM sodium azide, and then incubated with 1:20 dilution of appropriate FITC- or PE-labeled monoclonal antibodies for 30 minutes at 4 degrees C. After an additional wash, the labeled cells are analyzed by flow cytometry on a FAC Scan (Becton Dickinson).

Effect on the production of cytokines. Cytokines generated by dendritic cells, in particular IL-12, are important in the initiation of T-cell dependent immune responses. IL-12 strongly influences the development of Th1 helper T-cell immune response, and induces cytotoxic T and NK cell function. An ELISA is used to measure the IL-12 release as follows. Dendritic cells (10⁶/ml) are treated with increasing concentrations of agonists or antagonists of the invention for 24 hours. LPS (100 ng/ml) is added to the cell culture as positive control. Supernatants from the cell cultures are then collected and analyzed for IL-12 content using commercial ELISA kit (e.g., R & D Systems (Minneapolis, Minn.)). The standard protocols provided with the kits are used.

Effect on the expression of MHC Class II, costimulatory and adhesion molecules. Three major families of cell surface antigens can be identified on monocytes: adhesion molecules, molecules involved in antigen presentation, and Fc receptor. Modulation of the expression of MHC class II antigens and other costimulatory molecules, such as B7 and ICAM-1, may result in changes in the antigen presenting capacity of monocytes and ability to induce T cell activation. Increased expression of Fc receptors may correlate with improved monocyte cytotoxic activity, cytokine release and phagocytosis.

FACS analysis is used to examine the surface antigens as follows. Monocytes are treated 1-5 days with increasing concentrations of agonists or antagonists of the invention or LPS (positive control), washed with PBS containing 1% BSA and 0.02 mM sodium azide, and then incubated with 1:20 dilution of appropriate FITC- or PE-labeled monoclonal antibodies for 30 minutes at 4 degrees C. After an additional wash, the labeled cells are analyzed by flow cytometry on a FACScan (Becton Dickinson).

Monocyte activation and/or increased survival. Assays for molecules that activate (or alternatively, inactivate) monocytes and/or increase monocyte survival (or alternatively, decrease monocyte survival) are known in the art and may routinely be applied to determine whether a molecule of the invention functions as an inhibitor or activator of monocytes. Agonists or antagonists of the invention can be screened using the three assays described below. For each of these assays, Peripheral blood mononuclear cells (PBMC) are purified from single donor leukopacks (American Red Cross, Baltimore, Md.) by centrifugation through a HISTOPAQUE™ gradient (Sigma). Monocytes are isolated from PBMC by counterflow centrifugal elutriation.

Monocyte Survival Assay. Human peripheral blood monocytes progressively lose viability when cultured in absence of serum or other stimuli. Their death results from internally regulated processes (apoptosis). Addition to the culture of activating factors, such as TNF-alpha dramatically improves cell survival and prevents DNA fragmentation. Propidium iodide (PI) staining is used to measure apoptosis as follows. Monocytes are cultured for 48 hours in polypropylene tubes in serum-free medium (positive control), in the presence of 100 ng/ml TNF-alpha (negative control), and in the presence of varying concentrations of the compound to be tested. Cells are suspended at a concentration of 2×10⁶/ml in PBS containing PI at a final concentration of 5 μg/ml, and then incubated at room temperature for 5 minutes before FACScan analysis. PI uptake has been demonstrated to correlate with DNA fragmentation in this experimental paradigm.

Effect on cytokine release. An important function of monocytes/macrophages is their regulatory activity on other cellular populations of the immune system through the release of cytokines after stimulation. An ELISA to measure cytokine release is performed as follows. Human monocytes are incubated at a density of 5×10⁵ cells/ml with increasing concentrations of agonists or antagonists of the invention and under the same conditions, but in the absence of agonists or antagonists. For IL-12 production, the cells are primed overnight with IFN (100 U/ml) in the presence of agonist or antagonist of the invention. LPS (10 ng/ml) is then added. Conditioned media are collected after 24 h and kept frozen until use. Measurement of TNF-alpha, IL-10, MCP-1 and IL-8 is then performed using a commercially available ELISA kit (e.g., R & D Systems (Minneapolis, Minn.)) and applying the standard protocols provided with the kit.

Oxidative burst. Purified monocytes are plated in 96-w plate at 2-1×10⁵ cell/well. Increasing concentrations of agonists or antagonists of the invention are added to the wells in a total volume of 0.2 ml culture medium (RPMI 1640 ±10% FCS, glutamine and antibiotics). After 3 days incubation, the plates are centrifuged and the medium is removed from the wells. To the macrophage monolayers, 0.2 ml per well of phenol red solution (140 mM NaCl, 10 mM potassium phosphate buffer pH 7.0, 5.5 mM dextrose, 0.56 mM phenol red and 19 U/ml of HRPO) is added, together with the stimulant (200 nM PMA). The plates are incubated at 37° C. for 2 hours and the reaction is stopped by adding 20 μl 1N NaOH per well. The absorbance is read at 610 nm. To calculate the amount of H₂O₂ produced by the macrophages, a standard curve of a H₂O₂ solution of known molarity is performed for each experiment.

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 24 Biological Effects of Agonists or Antagonists of the Invention Astrocyte and Neuronal Assays'

Agonists or antagonists of the invention, expressed in Escherichia coli and purified as described above, can be tested for activity in promoting the survival, neurite outgrowth, or phenotypic differentiation of cortical neuronal cells and for inducing the proliferation of glial fibrillary acidic protein immunopositive cells, astrocytes. The selection of cortical cells for the bioassay is based on the prevalent expression of FGF-1 and FGF-2 in cortical structures and on the previously reported enhancement of cortical neuronal survival resulting from FGF-2 treatment. A thymidine incorporation assay, for example, can be used to elucidate an agonist or antagonist of the invention's activity on these cells.

Moreover, previous reports describing the biological effects of FGF-2 (basic FGF) on cortical or hippocampal neurons in vitro have demonstrated increases in both neuron survival and neurite outgrowth (Walicke et al., “Fibroblast growth factor promotes survival of dissociated hippocampal neurons and enhances neurite extension.” Proc. Natl. Acad. Sci. USA 83:3012-3016. (1986), assay herein incorporated by reference in its entirety). However, reports from experiments done on PC-12 cells suggest that these two responses are not necessarily synonymous and may depend on not only which FGF is being tested but also on which receptor(s) are expressed on the target cells. Using the primary cortical neuronal culture paradigm, the ability of an agonist or antagonist of the invention to induce neurite outgrowth can be compared to the response achieved with FGF-2 using, for example, a thymidine incorporation assay.

Fibroblast and Endothelial Cell Assays.

Human lung fibroblasts are obtained from Clonetics (San Diego, Calif.) and maintained in growth media from Clonetics. Dermal microvascular endothelial cells are obtained from Cell Applications (San Diego, Calif.). For proliferation assays, the human lung fibroblasts and dermal microvascular endothelial cells can be cultured at 5,000 cells/well in a 96-well plate for one day in growth medium. The cells are then incubated for one day in 0.1% BSA basal medium. After replacing the medium with fresh 0.1% BSA medium, the cells are incubated with the test proteins for 3 days. ALAMAR BLUE™ (Alamar Biosciences, Sacramento, Calif.) is added to each well to a final concentration of 10%. The cells are incubated for 4 hr. Cell viability is measured by reading in a CYTOFLUOR™ fluorescence reader. For the PGE₂ assays, the human lung fibroblasts are cultured at 5,000 cells/well in a 96-well plate for one day. After a medium change to 0.1% BSA basal medium, the cells are incubated with FGF-2 or agonists or antagonists of the invention with or without IL-1α for 24 hours. The supernatants are collected and assayed for PGE₂ by EIA kit (Cayman, Ann Arbor, Mich.). For the IL-6 assays, the human lung fibroblasts are cultured at 5,000 cells/well in a 96-well plate for one day. After a medium change to 0.1% BSA basal medium, the cells are incubated with FGF-2 or with or without agonists or antagonists of the invention IL-1α for 24 hours. The supernatants are collected and assayed for IL-6 by ELISA kit (Endogen, Cambridge, Mass.).

Human lung fibroblasts are cultured with FGF-2 or agonists or antagonists of the invention for 3 days in basal medium before the addition of ALAMAR BLUE™ to assess effects on growth of the fibroblasts. FGF-2 should show a stimulation at 10-2500 ng/ml which can be used to compare stimulation with agonists or antagonists of the invention.

Parkinson Models.

The loss of motor function in Parkinson's disease is attributed to a deficiency of striatal dopamine resulting from the degeneration of the nigrostriatal dopaminergic projection neurons. An animal model for Parkinson's that has been extensively characterized involves the systemic administration of 1-methyl-4 phenyl 1,2,3,6-tetrahydropyridine (MPTP). In the CNS, MPTP is taken-up by astrocytes and catabolized by monoamine oxidase B to 1-methyl-4-phenyl pyridine (MPP⁺) and released. Subsequently, MPP⁺ is actively accumulated in dopaminergic neurons by the high-affinity reuptake transporter for dopamine. MPP⁺ is then concentrated in mitochondria by the electrochemical gradient and selectively inhibits nicotidamide adenine disphosphate: ubiquinone oxidoreductionase (complex I), thereby interfering with electron transport and eventually generating oxygen radicals.

It has been demonstrated in tissue culture paradigms that FGF-2 (basic FGF) has trophic activity towards nigral dopaminergic neurons (Ferrari et al., Dev. Biol. 1989). Recently, Dr. Unsicker's group has demonstrated that administering FGF-2 in gel foam implants in the striatum results in the near complete protection of nigral dopaminergic neurons from the toxicity associated with MPTP exposure (Otto and Unsicker, J. Neuroscience, 1990).

Based on the data with FGF-2, agonists or antagonists of the invention can be evaluated to determine whether it has an action similar to that of FGF-2 in enhancing dopaminergic neuronal survival in vitro and it can also be tested in vivo for protection of dopaminergic neurons in the striatum from the damage associated with MPTP treatment. The potential effect of an agonist or antagonist of the invention is first examined in vitro in a dopaminergic neuronal cell culture paradigm. The cultures are prepared by dissecting the midbrain floor plate from gestation day 14 Wistar rat embryos. The tissue is dissociated with trypsin and seeded at a density of 200,000 cells/cm² on polyorthinine-laminin coated glass coverslips. The cells are maintained in Dulbecco's Modified Eagle's medium and F12 medium containing hormonal supplements (N1). The cultures are fixed with paraformaldehyde after 8 days in vitro and are processed for tyrosine hydroxylase, a specific marker for dopaminergic neurons, immunohistochemical staining. Dissociated cell cultures are prepared from embryonic rats. The culture medium is changed every third day and the factors are also added at that time.

Since the dopaminergic neurons are isolated from animals at gestation day 14, a developmental time which is past the stage when the dopaminergic precursor cells are proliferating, an increase in the number of tyrosine hydroxylase immunopositive neurons would represent an increase in the number of dopaminergic neurons surviving in vitro. Therefore, if an agonist or antagonist of the invention acts to prolong the survival of dopaminergic neurons, it would suggest that the agonist or antagonist may be involved in Parkinson's Disease.

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 25 The Effect of Agonists or Antagonists of the Invention on the Growth of Vascular Endothelial Cells

On day 1, human umbilical vein endothelial cells (HUVEC) are seeded at 2-5×10⁴ cells/35 mm dish density in M199 medium containing 4% fetal bovine serum (FBS), 16 units/ml heparin, and 50 units/ml endothelial cell growth supplements (ECGS, Biotechnique, Inc.). On day 2, the medium is replaced with M199 containing 10% FBS, 8 units/ml heparin. An agonist or antagonist of the invention, and positive controls, such as VEGF and basic FGF (bFGF) are added, at varying concentrations. On days 4 and 6, the medium is replaced. On day 8, cell number is determined with a Coulter Counter.

An increase in the number of HUVEC cells indicates that the compound of the invention may proliferate vascular endothelial cells, while a decrease in the number of HUVEC cells indicates that the compound of the invention inhibits vascular endothelial cells.

The studies described in this example tested activity of a polypeptide of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), agonists, and/or antagonists of the invention.

Example 26 Rat Corneal Wound Healing Model

This animal model shows the effect of an agonist or antagonist of the invention on neovascularization. The experimental protocol includes:

a) Making a 1-1.5 mm long incision from the center of cornea into the stromal layer. b) Inserting a spatula below the lip of the incision facing the outer corner of the eye. c) Making a pocket (its base is 1-1.5 mm form the edge of the eye). d) Positioning a pellet, containing 50 ng-5 μg of an agonist or antagonist of the invention, within the pocket. e) Treatment with an agonist or antagonist of the invention can also be applied topically to the corneal wounds in a dosage range of 20 mg-500 mg (daily treatment for five days).

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 27 Diabetic Mouse and Glucocorticoid-Impaired Wound Healing Models

Diabetic db+/db+ Mouse Model.

To demonstrate that an agonist or antagonist of the invention accelerates the healing process, the genetically diabetic mouse model of wound healing is used. The full thickness wound healing model in the db+/db+ mouse is a well characterized, clinically relevant and reproducible model of impaired wound healing. Healing of the diabetic wound is dependent on formation of granulation tissue and re-epithelialization rather than contraction (Gartner, M. H. et al., J. Surg. Res. 52:389 (1992); Greenhalgh, D. G. et al., Am. J. Pathol. 136:1235 (1990)).

The diabetic animals have many of the characteristic features observed in Type II diabetes mellitus. Homozygous (db+/db+) mice are obese in comparison to their normal heterozygous (db+/+m) littermates. Mutant diabetic (db+/db+) mice have a single autosomal recessive mutation on chromosome 4 (db+) (Coleman et al. Proc. Natl. Acad. Sci. USA 77:283-293 (1982)). Animals show polyphagia, polydipsia and polyuria. Mutant diabetic mice (db+/db+) have elevated blood glucose, increased or normal insulin levels, and suppressed cell-mediated immunity (Mandel et al., J. Immunol. 120:1375 (1978); Debray-Sachs, M. et al., Clin. Exp. Immunol. 51(1):1-7 (1983); Leiter et al., Am. J. of Pathol. 114:46-55 (1985)). Peripheral neuropathy, myocardial complications, and microvascular lesions, basement membrane thickening and glomerular filtration abnormalities have been described in these animals (Norido, F. et al., Exp. Neurol. 83(2):221-232 (1984); Robertson et al., Diabetes 29(1):60-67 (1980); Giacomelli et al., Lab Invest. 40(4):460-473 (1979); Coleman, D. L., Diabetes 31 (Suppl):1-6 (1982)). These homozygous diabetic mice develop hyperglycemia that is resistant to insulin analogous to human type II diabetes (Mandel et al., J. Immunol. 120:1375-1377 (1978)).

The characteristics observed in these animals suggests that healing in this model may be similar to the healing observed in human diabetes (Greenhalgh, et al., Am. J. of Pathol. 136:1235-1246 (1990)).

Genetically diabetic female C57BL/KsJ (db+/db+) mice and their non-diabetic (db+/+m) heterozygous littermates are used in this study (Jackson Laboratories). The animals are purchased at 6 weeks of age and are 8 weeks old at the beginning of the study. Animals are individually housed and received food and water ad libitum. All manipulations are performed using aseptic techniques. The experiments are conducted according to the rules and guidelines of Human Genome Sciences, Inc. Institutional Animal Care and Use Committee and the Guidelines for the Care and Use of Laboratory Animals.

Wounding protocol is performed according to previously reported methods (Tsuboi, R. and Rifkin, D. B., J. Exp. Med. 172:245-251 (1990)). Briefly, on the day of wounding, animals are anesthetized with an intraperitoneal injection of Avertin (0.01 mg/mL), 2,2,2-tribromoethanol and 2-methyl-2-butanol dissolved in deionized water. The dorsal region of the animal is shaved and the skin washed with 70% ethanol solution and iodine. The surgical area is dried with sterile gauze prior to wounding. An 8 mm full-thickness wound is then created using a Keyes tissue punch. Immediately following wounding, the surrounding skin is gently stretched to eliminate wound expansion. The wounds are left open for the duration of the experiment. Application of the treatment is given topically for 5 consecutive days commencing on the day of wounding. Prior to treatment, wounds are gently cleansed with sterile saline and gauze sponges.

Wounds are visually examined and photographed at a fixed distance at the day of surgery and at two day intervals thereafter. Wound closure is determined by daily measurement on days 1-5 and on day 8. Wounds are measured horizontally and vertically using a calibrated Jameson caliper. Wounds are considered healed if granulation tissue is no longer visible and the wound is covered by a continuous epithelium.

An agonist or antagonist of the invention is administered using at a range different doses, from 4 mg to 500 mg per wound per day for 8 days in vehicle. Vehicle control groups received 50 mL of vehicle solution.

Animals are euthanized on day 8 with an intraperitoneal injection of sodium pentobarbital (300 mg/kg). The wounds and surrounding skin are then harvested for histology and immunohistochemistry. Tissue specimens are placed in 10% neutral buffered formalin in tissue cassettes between biopsy sponges for further processing.

Three groups of 10 animals each (5 diabetic and 5 non-diabetic controls) are evaluated: 1) Vehicle placebo control, 2) untreated group, and 3) treated group.

Wound closure is analyzed by measuring the area in the vertical and horizontal axis and obtaining the total square area of the wound. Contraction is then estimated by establishing the differences between the initial wound area (day 0) and that of post treatment (day 8). The wound area on day 1 is 64 mm², the corresponding size of the dermal punch. Calculations are made using the following formula:

[Open area on day 8]−[Open area on day 1]/[Open area on day 1]

Specimens are fixed in 10% buffered formalin and paraffin embedded blocks are sectioned perpendicular to the wound surface (5 mm) and cut using a Reichert-Jung microtome. Routine hematoxylin-eosin (H&E) staining is performed on cross-sections of bisected wounds. Histologic examination of the wounds are used to assess whether the healing process and the morphologic appearance of the repaired skin is altered by treatment with an agonist or antagonist of the invention. This assessment included verification of the presence of cell accumulation, inflammatory cells, capillaries, fibroblasts, re-epithelialization and epidermal maturity (Greenhalgh, D. G. et al., Am. J. Pathol. 136:1235 (1990)). A calibrated lens micrometer is used by a blinded observer.

Tissue sections are also stained immunohistochemically with a polyclonal rabbit anti-human keratin antibody using ABC Elite detection system. Human skin is used as a positive tissue control while non-immune IgG is used as a negative control. Keratinocyte growth is determined by evaluating the extent of reepithelialization of the wound using a calibrated lens micrometer.

Proliferating cell nuclear antigen/cyclin (PCNA) in skin specimens is demonstrated by using anti-PCNA antibody (1:50) with an ABC Elite detection system. Human colon cancer served as a positive tissue control and human brain tissue is used as a negative tissue control. Each specimen included a section with omission of the primary antibody and substitution with non-immune mouse IgG. Ranking of these sections is based on the extent of proliferation on a scale of 0-8, the lower side of the scale reflecting slight proliferation to the higher side reflecting intense proliferation.

Experimental data are analyzed using an unpaired t test. A p value of <0.05 is considered significant.

Steroid Impaired Rat Model

The inhibition of wound healing by steroids has been well documented in various in vitro and in vivo systems (Wahl, Glucocorticoids and Wound healing. In: Anti-Inflammatory Steroid Action: Basic and Clinical Aspects. 280-302 (1989); Wahl et al., J. Immunol. 115: 476-481 (1975); Werb et al., J. Exp. Med. 147:1684-1694 (1978)). Glucocorticoids retard wound healing by inhibiting angiogenesis, decreasing vascular permeability (Ebert et al., An. Intern. Med. 37:701-705 (1952)), fibroblast proliferation, and collagen synthesis (Beck et al., Growth Factors. 5: 295-304 (1991); Haynes et al., J. Clin. Invest. 61: 703-797 (1978)) and producing a transient reduction of circulating monocytes (Haynes et al., J. Clin. Invest. 61: 703-797 (1978); Wahl, “Glucocorticoids and wound healing”, In: Antiinflammatory Steroid Action: Basic and Clinical Aspects, Academic Press, New York, pp. 280-302 (1989)). The systemic administration of steroids to impaired wound healing is a well establish phenomenon in rats (Beck et al., Growth Factors. 5: 295-304 (1991); Haynes et al., J. Clin. Invest. 61: 703-797 (1978); Wahl, “Glucocorticoids and wound healing”, In: Antiinflammatory Steroid Action: Basic and Clinical Aspects, Academic Press, New York, pp. 280-302 (1989); Pierce et al., Proc. Natl. Acad. Sci. USA 86: 2229-2233 (1989)).

To demonstrate that an agonist or antagonist of the invention can accelerate the healing process, the effects of multiple topical applications of the agonist or antagonist on full thickness excisional skin wounds in rats in which healing has been impaired by the systemic administration of methylprednisolone is assessed.

Young adult male Sprague Dawley rats weighing 250-300 g (Charles River Laboratories) are used in this example. The animals are purchased at 8 weeks of age and are 9 weeks old at the beginning of the study. The healing response of rats is impaired by the systemic administration of methylprednisolone (17 mg/kg/rat intramuscularly) at the time of wounding. Animals are individually housed and received food and water ad libitum. All manipulations are performed using aseptic techniques. This study is conducted according to the rules and guidelines of Human Genome Sciences, Inc. Institutional Animal Care and Use Committee and the Guidelines for the Care and Use of Laboratory Animals.

The wounding protocol is followed according to section A, above. On the day of wounding, animals are anesthetized with an intramuscular injection of ketamine (50 mg/kg) and xylazine (5 mg/kg). The dorsal region of the animal is shaved and the skin washed with 70% ethanol and iodine solutions. The surgical area is dried with sterile gauze prior to wounding. An 8 mm full-thickness wound is created using a Keyes tissue punch. The wounds are left open for the duration of the experiment. Applications of the testing materials are given topically once a day for 7 consecutive days commencing on the day of wounding and subsequent to methylprednisolone administration. Prior to treatment, wounds are gently cleansed with sterile saline and gauze sponges.

Wounds are visually examined and photographed at a fixed distance at the day of wounding and at the end of treatment. Wound closure is determined by daily measurement on days 1-5 and on day 8. Wounds are measured horizontally and vertically using a calibrated Jameson caliper. Wounds are considered healed if granulation tissue is no longer visible and the wound is covered by a continuous epithelium.

The agonist or antagonist of the invention is administered using at a range different doses, from 4 mg to 500 mg per wound per day for 8 days in vehicle. Vehicle control groups received 50 mL of vehicle solution.

Animals are euthanized on day 8 with an intraperitoneal injection of sodium pentobarbital (300 mg/kg). The wounds and surrounding skin are then harvested for histology. Tissue specimens are placed in 10% neutral buffered formalin in tissue cassettes between biopsy sponges for further processing.

Three groups of 10 animals each (5 with methylprednisolone and 5 without glucocorticoid) are evaluated: 1) Untreated group 2) Vehicle placebo control 3) treated groups.

Wound closure is analyzed by measuring the area in the vertical and horizontal axis and obtaining the total area of the wound. Closure is then estimated by establishing the differences between the initial wound area (day 0) and that of post treatment (day 8). The wound area on day 1 is 64 mm², the corresponding size of the dermal punch. Calculations are made using the following formula:

[Open area on day 8]−[Open area on day 1]/[Open area on day 1]

Specimens are fixed in 10% buffered formalin and paraffin embedded blocks are sectioned perpendicular to the wound surface (5 mm) and cut using an Olympus microtome. Routine hematoxylin-eosin (H&E) staining is performed on cross-sections of bisected wounds. Histologic examination of the wounds allows assessment of whether the healing process and the morphologic appearance of the repaired skin is improved by treatment with an agonist or antagonist of the invention. A calibrated lens micrometer is used by a blinded observer to determine the distance of the wound gap.

Experimental data are analyzed using an unpaired t test. A p value of <0.05 is considered significant.

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 28 Lymphadema Animal Model

The purpose of this experimental approach is to create an appropriate and consistent lymphedema model for testing the therapeutic effects of an agonist or antagonist of the invention in lymphangiogenesis and re-establishment of the lymphatic circulatory system in the rat hind limb. Effectiveness is measured by swelling volume of the affected limb, quantification of the amount of lymphatic vasculature, total blood plasma protein, and histopathology. Acute lymphedema is observed for 7-10 days. Perhaps more importantly, the chronic progress of the edema is followed for up to 3-4 weeks.

Prior to beginning surgery, blood sample is drawn for protein concentration analysis. Male rats weighing approximately ˜350 g are dosed with Pentobarbital. Subsequently, the right legs are shaved from knee to hip. The shaved area is swabbed with gauze soaked in 70% EtOH. Blood is drawn for serum total protein testing. Circumference and volumetric measurements are made prior to injecting dye into paws after marking 2 measurement levels (0.5 cm above heel, at mid-pt of dorsal paw). The intradermal dorsum of both right and left paws are injected with 0.05 ml of 1% Evan's Blue. Circumference and volumetric measurements are then made following injection of dye into paws.

Using the knee joint as a landmark, a mid-leg inguinal incision is made circumferentially allowing the femoral vessels to be located. Forceps and hemostats are used to dissect and separate the skin flaps. After locating the femoral vessels, the lymphatic vessel that runs along side and underneath the vessel(s) is located. The main lymphatic vessels in this area are then electrically coagulated or suture ligated.

Using a microscope, muscles in back of the leg (near the semitendinosis and adductors) are bluntly dissected. The popliteal lymph node is then located. The 2 proximal and 2 distal lymphatic vessels and distal blood supply of the popliteal node are then ligated by suturing. The popliteal lymph node, and any accompanying adipose tissue, is then removed by cutting connective tissues.

Care is taken to control any mild bleeding resulting from this procedure. After lymphatics are occluded, the skin flaps are sealed by using liquid skin (Vetbond) (AJ Buck). The separated skin edges are sealed to the underlying muscle tissue while leaving a gap of ˜0.5 cm around the leg. Skin also may be anchored by suturing to underlying muscle when necessary.

To avoid infection, animals are housed individually with mesh (no bedding). Recovering animals are checked daily through the optimal edematous peak, which typically occurred by day 5-7. The plateau edematous peak are then observed. To evaluate the intensity of the lymphedema, the circumference and volumes of 2 designated places on each paw before operation and daily for 7 days are measured. The effect of plasma proteins on lymphedema is determined and whether protein analysis is a useful testing perimeter is also investigated. The weights of both control and edematous limbs are evaluated at 2 places. Analysis is performed in a blind manner.

Circumference Measurements: Under brief gas anesthetic to prevent limb movement, a cloth tape is used to measure limb circumference. Measurements are done at the ankle bone and dorsal paw by 2 different people and those 2 readings are averaged. Readings are taken from both control and edematous limbs.

Volumetric Measurements: On the day of surgery, animals are anesthetized with Pentobarbital and are tested prior to surgery. For daily volumetrics animals are under brief halothane anesthetic (rapid immobilization and quick recovery), and both legs are shaved and equally marked using waterproof marker on legs. Legs are first dipped in water, then dipped into instrument to each marked level then measured by Buxco edema software (Chen/Victor). Data is recorded by one person, while the other is dipping the limb to marked area.

Blood-plasma protein measurements: Blood is drawn, spun, and serum separated prior to surgery and then at conclusion for total protein and Ca2⁺ comparison.

Limb Weight Comparison: After drawing blood, the animal is prepared for tissue collection. The limbs are amputated using a quillitine, then both experimental and control legs are cut at the ligature and weighed. A second weighing is done as the tibio-cacaneal joint is disarticulated and the foot is weighed.

Histological Preparations: The transverse muscle located behind the knee (popliteal) area is dissected and arranged in a metal mold, filled with freezeGel, dipped into cold methylbutane, placed into labeled sample bags at −80EC until sectioning. Upon sectioning, the muscle is observed under fluorescent microscopy for lymphatics.

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 29 Suppression of TNF Alpha-induced Adhesion Molecule Expression by an Agonist or Antagonist of the Invention

The recruitment of lymphocytes to areas of inflammation and angiogenesis involves specific receptor-ligand interactions between cell surface adhesion molecules (CAMs) on lymphocytes and the vascular endothelium. The adhesion process, in both normal and pathological settings, follows a multi-step cascade that involves intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and endothelial leukocyte adhesion molecule-1 (E-selectin) expression on endothelial cells (EC). The expression of these molecules and others on the vascular endothelium determines the efficiency with which leukocytes may adhere to the local vasculature and extravasate into the local tissue during the development of an inflammatory response. The local concentration of cytokines and growth factor participate in the modulation of the expression of these CAMs.

Tumor necrosis factor alpha (TNF-a), a potent proinflammatory cytokine, is a stimulator of all three CAMs on endothelial cells and may be involved in a wide variety of inflammatory responses, often resulting in a pathological outcome.

The potential of an agonist or antagonist of the invention to mediate a suppression of TNF-a induced CAM expression can be examined. A modified ELISA assay which uses ECs as a solid phase absorbent is employed to measure the amount of CAM expression on TNF-a treated ECs when co-stimulated with a member of the FGF family of proteins.

To perform the experiment, human umbilical vein endothelial cell (HUVEC) cultures are obtained from pooled cord harvests and maintained in growth medium (EGM-2; Clonetics, San Diego, Calif.) supplemented with 10% FCS and 1% penicillin/streptomycin in a 37 degree C. humidified incubator containing 5% CO₂. HUVECs are seeded in 96-well plates at concentrations of 1×10⁴ cells/well in EGM medium at 37 degree C. for 18-24 hrs or until confluent. The monolayers are subsequently washed 3 times with a serum-free solution of RPMI-1640 supplemented with 100 U/ml penicillin and 100 mg/ml streptomycin, and treated with a given cytokine and/or growth factor(s) for 24 h at 37 degree C. Following incubation, the cells are then evaluated for CAM expression.

Human Umbilical Vein Endothelial cells (HUVECs) are grown in a standard 96 well plate to confluence. Growth medium is removed from the cells and replaced with 90 μl of 199 Medium (10% FBS). Samples for testing and positive or negative controls are added to the plate in triplicate (in 10 μl volumes). Plates are incubated at 37 degree C. for either 5 h (selectin and integrin expression) or 24 h (integrin expression only). Plates are aspirated to remove medium and 100 μl of 0.1% paraformaldehyde-PBS (with Ca++ and Mg++) is added to each well. Plates are held at 4° C. for 30 min.

Fixative is then removed from the wells and wells are washed 1× with PBS(+Ca,Mg)+0.5% BSA and drained. Do not allow the wells to dry. Add 10 μl of diluted primary antibody to the test and control wells. Anti-ICAM-1-Biotin, Anti-VCAM-1-Biotin and Anti-E-selectin-Biotin are used at a concentration of 10 μg/ml (1:10 dilution of 0.1 mg/ml stock antibody). Cells are incubated at 37° C. for 30 min. in a humidified environment. Wells are washed ×3 with PBS(+Ca,Mg)+0.5% BSA.

Then add 20 μl of diluted ExtrAvidin-Alkaline Phosphotase (1:5,000 dilution) to each well and incubated at 37° C. for 30 min. Wells are washed ×3 with PBS(+Ca,Mg)+0.5% BSA. 1 tablet of p-Nitrophenol Phosphate pNPP is dissolved in 5 ml of glycine buffer (pH 10.4). 100 μl of pNPP substrate in glycine buffer is added to each test well. Standard wells in triplicate are prepared from the working dilution of the ExtrAvidin-Alkaline Phosphotase in glycine buffer: 1:5,000 (10⁰)>10^(−0.5)>10⁻¹>10^(−1.5).5 μl of each dilution is added to triplicate wells and the resulting AP content in each well is 5.50 ng, 1.74 ng, 0.55 ng, 0.18 ng. 100 μl of pNNP reagent must then be added to each of the standard wells. The plate must be incubated at 37° C. for 4 h. A volume of 50 μl of 3M NaOH is added to all wells. The results are quantified on a plate reader at 405 nm. The background subtraction option is used on blank wells filled with glycine buffer only. The template is set up to indicate the concentration of AP-conjugate in each standard well [5.50 ng; 1.74 ng; 0.55 ng; 0.18 ng]. Results are indicated as amount of bound AP-conjugate in each sample.

The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 30 Production of Polypeptide of the Invention For High-Throughput Screening Assays

The following protocol produces a supernatant containing polypeptide of the present invention to be tested. This supernatant can then be used in the Screening Assays described in Examples 32-41.

First, dilute Poly-D-Lysine (644 587 Boehringer-Mannheim) stock solution (1 mg/ml in PBS) 1:20 in PBS (w/o calcium or magnesium 17-516F Biowhittaker) for a working solution of 50 ug/ml. Add 200 μl of this solution to each well (24 well plates) and incubate at RT for 20 minutes. Be sure to distribute the solution over each well (note: a 12-channel pipetter may be used with tips on every other channel). Aspirate off the Poly-D-Lysine solution and rinse with 1 ml PBS (Phosphate Buffered Saline). The PBS should remain in the well until just prior to plating the cells and plates may be poly-lysine coated in advance for up to two weeks.

Plate 293T cells (do not carry cells past P+20) at 2×10⁵ cells/well in 0.5 ml DMEM(Dulbecco's Modified Eagle Medium)(with 4.5 G/L glucose and L-glutamine (12-604F Biowhittaker))/10% heat inactivated FBS(14-503F Biowhittaker)/1×Penstrep(17-602E Biowhittaker). Let the cells grow overnight.

The next day, mix together in a sterile solution basin: 300 μl LIPOFECTAMINE™ (18324-012 Gibco/BRL) and 5 ml OPTI-MEM™ I (31985070 Gibco/BRL)/96-well plate. With a small volume multi-channel pipetter, aliquot approximately 2 μg of an expression vector containing a polynucleotide insert, produced by the methods described in Examples 8-10, into an appropriately labeled 96-well round bottom plate. With a multi-channel pipetter, add 50 μl of the LIPOFECTAMINE™/OPTI-MEM™ I mixture to each well. Pipette up and down gently to mix. Incubate at RT 15-45 minutes. After about 20 minutes, use a multi-channel pipetter to add 150 μl OPTI-MEM™ I to each well. As a control, one plate of vector DNA lacking an insert should be transfected with each set of transfections.

Preferably, the transfection should be performed by tag-teaming the following tasks. By tag-teaming, hands on time is cut in half, and the cells do not spend too much time on PBS. First, person A aspirates off the media from four 24-well plates of cells, and then person B rinses each well with 0.5-1 ml PBS. Person A then aspirates off PBS rinse, and person B, using a12-channel pipetter with tips on every other channel, adds the 200 μl of DNA/LIPOFECTAMINE™/OPTI-MEM™ I complex to the odd wells first, then to the even wells, to each row on the 24-well plates. Incubate at 37 degree C. for 6 hours.

While cells are incubating, prepare appropriate media, either 1% BSA in DMEM with 1×penstrep, or HGS CHO-5 media (116.6 mg/L of CaCl₂ (anhyd); 0.00130 mg/L CuSO₄.5H₂O; 0.050 mg/L of Fe(NO₃)₃-9H₂O; 0.417 mg/L of FeSO₄.7H₂O; 311.80 mg/L of Kcl; 28.64 mg/L of MgCl₂; 48.84 mg/L of MgSO₄; 6995.50 mg/L of NaCl; 2400.0 mg/L of NaHCO₃; 62.50 mg/L of NaH₂PO₄—H₂O; 71.02 mg/L of Na₂HPO₄; .4320 mg/L of ZnSO₄-7H₂O; 0.002 mg/L of Arachidonic Acid; 1.022 mg/L of Cholesterol; 0.070 mg/L of DL-alpha-Tocopherol-Acetate; 0.0520 mg/L of Linoleic Acid; 0.010 mg/L of Linolenic Acid; 0.010 mg/L of Myristic Acid; 0.010 mg/L of Oleic Acid; 0.010 mg/L of Palmitric Acid; 0.010 mg/L of Palmitic Acid; 100 mg/L of Pluronic F-68; 0.010 mg/L of Stearic Acid; 2.20 mg/L of Tween 80; 4551 mg/L of D-Glucose; 130.85 mg/ml of L-Alanine; 147.50 mg/ml of L-Arginine-HCL; 7.50 mg/ml of L-Asparagine-H₂O; 6.65 mg/ml of L-Aspartic Acid; 29.56 mg/ml of L-Cystine-2HCL-H₂O; 31.29 mg/ml of L-Cystine-2HCL; 7.35 mg/ml of L-Glutamic Acid; 365.0 mg/ml of L-Glutamine; 18.75 mg/ml of Glycine; 52.48 mg/ml of L-Histidine-HCL-H₂O; 106.97 mg/ml of L-Isoleucine; 111.45 mg/ml of L-Leucine; 163.75 mg/ml of L-Lysine HCL; 32.34 mg/ml of L-Methionine; 68.48 mg/ml of L-Phenylalainine; 40.0 mg/ml of L-Proline; 26.25 mg/ml of L-Serine; 101.05 mg/ml of L-Threonine; 19.22 mg/ml of L-Tryptophan; 91.79 mg/ml of L-Tryrosine-2Na-2H₂O; and 99.65 mg/ml of L-Valine; 0.0035 mg/L of Biotin; 3.24 mg/L of D-Ca Pantothenate; 11.78 mg/L of Choline Chloride; 4.65 mg/L of Folic Acid; 15.60 mg/L of i-Inositol; 3.02 mg/L of Niacinamide; 3.00 mg/L of Pyridoxal HCL; 0.031 mg/L of Pyridoxine HCL; 0.319 mg/L of Riboflavin; 3.17 mg/L of Thiamine HCL; 0.365 mg/L of Thymidine; 0.680 mg/L of Vitamin B₁₂; 25 mM of HEPES Buffer; 2.39 mg/L of Na Hypoxanthine; 0.105 mg/L of Lipoic Acid; 0.081 mg/L of Sodium Putrescine-2HCL; 55.0 mg/L of Sodium Pyruvate; 0.0067 mg/L of Sodium Selenite; 20 μM of Ethanolamine; 0.122 mg/L of Ferric Citrate; 41.70 mg/L of Methyl-B-Cyclodextrin complexed with Linoleic Acid; 33.33 mg/L of Methyl-B-Cyclodextrin complexed with Oleic Acid; 10 mg/L of Methyl-B-Cyclodextrin complexed with Retinal Acetate. Adjust osmolarity to 327 mOsm) with 2 mm glutamine and 1×penstrep. (BSA (81-068-3 Bayer) 100 gm dissolved in 1 L DMEM for a 10% BSA stock solution). Filter the media and collect 50 μl for endotoxin assay in 15 ml polystyrene conical.

The transfection reaction is terminated, preferably by tag-teaming, at the end of the incubation period. Person A aspirates off the transfection media, while person B adds 1.5 ml appropriate media to each well. Incubate at 37 degree C. for 45 or 72 hours depending on the media used: 1% BSA for 45 hours or CHO-5 for 72 hours.

On day four, using a 300 μl multichannel pipetter, aliquot 600 μl in one 1 ml deep well plate and the remaining supernatant into a 2 ml deep well. The supernatants from each well can then be used in the assays described in Examples 32-39.

It is specifically understood that when activity is obtained in any of the assays described below using a supernatant, the activity originates from either the polypeptide of the present invention directly (e.g., as a secreted protein) or by polypeptide of the present invention inducing expression of other proteins, which are then secreted into the supernatant. Thus, the invention further provides a method of identifying the protein in the supernatant characterized by an activity in a particular assay.

Example 31 Construction of GAS Reporter Construct

One signal transduction pathway involved in the differentiation and proliferation of cells is called the Jaks-STATs pathway. Activated proteins in the Jaks-STATs pathway bind to gamma activation site “GAS” elements or interferon-sensitive responsive element (“ISRE”), located in the promoter of many genes. The binding of a protein to these elements alter the expression of the associated gene.

GAS and ISRE elements are recognized by a class of transcription factors called Signal Transducers and Activators of Transcription, or “STATs.” There are six members of the STATs family. Stat1 and Stat3 are present in many cell types, as is Stat2 (as response to IFN-alpha is widespread). Stat4 is more restricted and is not in many cell types though it has been found in T helper class I, cells after treatment with IL-12. Stat5 was originally called mammary growth factor, but has been found at higher concentrations in other cells including myeloid cells. It can be activated in tissue culture cells by many cytokines.

The STATs are activated to translocate from the cytoplasm to the nucleus upon tyrosine phosphorylation by a set of kinases known as the Janus Kinase (“Jaks”) family. Jaks represent a distinct family of soluble tyrosine kinases and include Tyk2, Jak1, Jak2, and Jak3. These kinases display significant sequence similarity and are generally catalytically inactive in resting cells.

The Jaks are activated by a wide range of receptors summarized in the Table below. (Adapted from review by Schidler and Darnell, Ann. Rev. Biochem. 64:621-51 (1995)). A cytokine receptor family, capable of activating Jaks, is divided into two groups: (a) Class 1 includes receptors for IL-2, IL-3, IL-4, IL-6, IL-7, IL-9, IL-11, IL-12, IL-15, Epo, PRL, GH, G-CSF, GM-CSF, LIF, CNTF, and thrombopoietin; and (b) Class 2 includes IFN-a, IFN-g, and IL-10. The Class 1 receptors share a conserved cysteine motif (a set of four conserved cysteines and one tryptophan) and a WSXWS motif (a membrane proximal region encoding Trp-Ser-Xaa-Trp-Ser (SEQ ID NO: 2)).

Thus, on binding of a ligand to a receptor, Jaks are activated, which in turn activate STATs, which then translocate and bind to GAS elements. This entire process is encompassed in the Jaks-STATs signal transduction pathway. Therefore, activation of the Jaks-STATs pathway, reflected by the binding of the GAS or the ISRE element, can be used to indicate proteins involved in the proliferation and differentiation of cells. For example, growth factors and cytokines are known to activate the Jaks-STATs pathway (See Table below). Thus, by using GAS elements linked to reporter molecules, activators of the Jaks-STATs pathway can be identified.

JAKs Ligand tyk2 Jak1 Jak2 Jak3 STATS GAS (elements) or ISRE IFN family IFN-a/B + + − − 1, 2, 3 ISRE IFN-g + + − 1 GAS (IRF1 > Lys6 > IFP) Il-10 + ? ? − 1, 3 gp130 family IL-6 (Pleiotropic) + + + ? 1, 3 GAS (IRF1 > Lys6 > IFP) Il-11 (Pleiotropic) ? + ? ? 1, 3 OnM (Pleiotropic) ? + + ? 1, 3 LIF (Pleiotropic) ? + + ? 1, 3 CNTF (Pleiotropic) −/+ + + ? 1, 3 G-CSF (Pleiotropic) ? + ? ? 1, 3 IL-12 (Pleiotropic) + − + + 1, 3 g-C family IL-2 (lymphocytes) − + − + 1, 3, 5 GAS IL-4 (lymph/myeloid) − + − + 6 GAS (IRF1 = IFP >> Ly6)(IgH) IL-7 (lymphocytes) − + − + 5 GAS IL-9 (lymphocytes) − + − + 5 GAS IL-13 (lymphocyte) − + ? ? 6 GAS IL-15 ? + ? + 5 GAS gp140 family IL-3 (myeloid) − − + − 5 GAS (IRF1 > IFP >> Ly6) IL-5 (myeloid) − − + − 5 GAS GM-CSF (myeloid) − − + − 5 GAS Growth hormone family GH ? − + − 5 PRL ? +/− + − 1, 3, 5 EPO ? − + − 5 GAS (B-CAS > IRF1 = IFP >> Ly6) Receptor Tyrosine Kinases EGF ? + + − 1, 3 GAS (IRF1) PDGF ? + + − 1, 3 CSF-1 ? + + − 1, 3 GAS (not IRF1)

To construct a synthetic GAS containing promoter element, which is used in the Biological Assays described in Examples 32-33, a PCR based strategy is employed to generate a GAS-SV40 promoter sequence. The 5′ primer contains four tandem copies of the GAS binding site found in the IRF1 promoter and previously demonstrated to bind STATs upon induction with a range of cytokines (Rothman et al., Immunity 1:457-468 (1994).), although other GAS or ISRE elements can be used instead. The 5′ primer also contains 18 bp of sequence complementary to the SV40 early promoter sequence and is flanked with an XhoI site. The sequence of the 5′ primer is:

(SEQ ID NO: 3) 5′: GCGCCTCGAGATTTCCCCGAAATCTAGATTTCCCCGAAATGATTTC CCCGAAATGATTTCCCCGAAATATCTGCCATCTCAATTAG: 3′

The downstream primer is complementary to the SV40 promoter and is flanked with a Hind III site:

(SEQ ID NO: 4) 5′: GCGGCAAGCTTTTTGCAAAGCCTAGGC: 3′

PCR amplification is performed using the SV40 promoter template present in the B-gal:promoter plasmid obtained from CLONTECH™. The resulting PCR fragment is digested with XhoI/Hind III and subcloned into BLSK2-. (STRATAGENE™.) Sequencing with forward and reverse primers confirms that the insert contains the following sequence:

(SEQ ID NO: 5) 5′: CTCGAGATTTCCCCGAAATCTAGATTTCCCCGAAATGATTTCCCCG AAATGATTTCCCCGAAATATCTGCCATCTCAATTAGTCAGCAACCATAGT CCCGCCCCTAACTCCGCCCATCCCGCCCCTAACTCCGCCCAGTTCCGCC CATTCTCCGCCCCATGGCTGACTAATTTTTTTTATTTATGCAGAGGCCGA GGCCGCCTCGGCCTCTGAGCTATTCCAGAAGTAGTGAGGAGGCTTTTTT GGAGGCCTAGGCTTTTGCAAAAAGCTT: 3′

With this GAS promoter element linked to the SV40 promoter, a GAS:SEAP2 reporter construct is next engineered. Here, the reporter molecule is a secreted alkaline phosphatase, or “SEAP.” Clearly, however, any reporter molecule can be instead of SEAP, in this or in any of the other Examples. Well known reporter molecules that can be used instead of SEAP include chloramphenicol acetyltransferase (CAT), luciferase, alkaline phosphatase, B-galactosidase, green fluorescent protein (GFP), or any protein detectable by an antibody.

The above sequence confirmed synthetic GAS-SV40 promoter element is subcloned into the pSEAP-Promoter vector obtained from CLONTECH™ using HindIII and XhoI, effectively replacing the SV40 promoter with the amplified GAS: SV40 promoter element, to create the GAS-SEAP vector. However, this vector does not contain a neomycin resistance gene, and therefore, is not preferred for mammalian expression systems.

Thus, in order to generate mammalian stable cell lines expressing the GAS-SEAP reporter, the GAS-SEAP cassette is removed from the GAS-SEAP vector using SalI and NotI, and inserted into a backbone vector containing the neomycin resistance gene, such as pGFP-1 (CLONTECH™), using these restriction sites in the multiple cloning site, to create the GAS-SEAP/Neo vector. Once this vector is transfected into mammalian cells, this vector can then be used as a reporter molecule for GAS binding as described in Examples 32-33.

Other constructs can be made using the above description and replacing GAS with a different promoter sequence. For example, construction of reporter molecules containing EGR and NF-KB promoter sequences are described in Examples 34 and 35. However, many other promoters can be substituted using the protocols described in these Examples. For instance, SRE, IL-2, NFAT, or Osteocalcin promoters can be substituted, alone or in combination (e.g., GAS/NF-KB/EGR, GAS/NF-KB, Il-2/NFAT, or NF-KB/GAS). Similarly, other cell lines can be used to test reporter construct activity, such as HELA (epithelial), HUVEC (endothelial), Reh (B-cell), Saos-2 (osteoblast), HUVAC (aortic), or Cardiomyocyte.

Example 32 High-Throughput Screening Assay for T-cell Activity

The following protocol is used to assess T-cell activity by identifying factors, and determining whether supernate containing a polypeptide of the invention proliferates and/or differentiates T-cells. T-cell activity is assessed using the GAS/SEAP/Neo construct produced in Example 31. Thus, factors that increase SEAP activity indicate the ability to activate the Jaks-STATS signal transduction pathway. The T-cell used in this assay is Jurkat T-cells (ATCC™ Accession No. TIB-152), although Molt-3 cells (ATCC™ Accession No. CRL-1552) and Molt-4 cells (ATCC™ Accession No. CRL-1582) cells can also be used.

Jurkat T-cells are lymphoblastic CD4+Th1 helper cells. In order to generate stable cell lines, approximately 2 million Jurkat cells are transfected with the GAS-SEAP/neo vector using DMRIE-C (Life Technologies)(transfection procedure described below). The transfected cells are seeded to a density of approximately 20,000 cells per well and transfectants resistant to 1 mg/ml genticin selected. Resistant colonies are expanded and then tested for their response to increasing concentrations of interferon gamma. The dose response of a selected clone is demonstrated.

Specifically, the following protocol will yield sufficient cells for 75 wells containing 200 μl of cells. Thus, it is either scaled up, or performed in multiple to generate sufficient cells for multiple 96 well plates. Jurkat cells are maintained in RPMI+10% serum with 1% Pen-Strep. Combine 2.5 mls of OPTI-MEM™ (Life Technologies) with 10 μg of plasmid DNA in a T25 flask. Add 2.5 ml OPTI-MEM™ containing 50 μl of DMRIE-C and incubate at room temperature for 15-45 mins.

During the incubation period, count cell concentration, spin down the required number of cells (10⁷ per transfection), and resuspend in OPTI-MEM™ to a final concentration of 10⁷ cells/ml. Then add 1 ml of 1×10⁷ cells in OPTI-MEM™ to T25 flask and incubate at 37 degree C. for 6 hrs. After the incubation, add 10 ml of RPMI+15% serum.

The Jurkat:GAS-SEAP stable reporter lines are maintained in RPMI+10% serum, 1 mg/ml Genticin, and 1% Pen-Strep. These cells are treated with supernatants containing polypeptide of the present invention or polypeptide of the present invention induced polypeptides as produced by the protocol described in Example 30.

On the day of treatment with the supernatant, the cells should be washed and resuspended in fresh RPMI+10% serum to a density of 500,000 cells per ml. The exact number of cells required will depend on the number of supernatants being screened. For one 96 well plate, approximately 10 million cells (for 10 plates, 100 million cells) are required.

Transfer the cells to a triangular reservoir boat, in order to dispense the cells into a 96 well dish, using a 12 channel pipette. Using a 12 channel pipette, transfer 200 μl of cells into each well (therefore adding 100,000 cells per well).

After all the plates have been seeded, 50 μl of the supernatants are transferred directly from the 96 well plate containing the supernatants into each well using a 12 channel pipette. In addition, a dose of exogenous interferon gamma (0.1, 1.0, 10 ng) is added to wells H9, H10, and H11 to serve as additional positive controls for the assay.

The 96 well dishes containing Jurkat cells treated with supernatants are placed in an incubator for 48 hrs (note: this time is variable between 48-72 hrs). 35 μl samples from each well are then transferred to an opaque 96 well plate using a 12 channel pipette. The opaque plates should be covered (using sellophene covers) and stored at −20 degree C. until SEAP assays are performed according to Example 36. The plates containing the remaining treated cells are placed at 4 degree C. and serve as a source of material for repeating the assay on a specific well if desired.

As a positive control, 100 Unit/ml interferon gamma can be used which is known to activate Jurkat T cells. Over 30 fold induction is typically observed in the positive control wells.

The above protocol may be used in the generation of both transient, as well as, stable transfected cells, which would be apparent to those of skill in the art.

Example 33 High-Throughput Screening Assay Identifying Myeloid Activity

The following protocol is used to assess myeloid activity of polypeptide of the present invention by determining whether polypeptide of the present invention proliferates and/or differentiates myeloid cells. Myeloid cell activity is assessed using the GAS/SEAP/Neo construct produced in Example 31. Thus, factors that increase SEAP activity indicate the ability to activate the Jaks-STATS signal transduction pathway. The myeloid cell used in this assay is U937, a pre-monocyte cell line, although TF-1, HL60, or KG1 can be used.

To transiently transfect U937 cells with the GAS/SEAP/Neo construct produced in Example 31, a DEAE-Dextran method (Kharbanda et. al., 1994, Cell Growth & Differentiation, 5:259-265) is used. First, harvest 2×10⁷ U937 cells and wash with PBS. The U937 cells are usually grown in RPMI 1640 medium containing 10% heat-inactivated fetal bovine serum (FBS) supplemented with 100 units/ml penicillin and 100 mg/ml streptomycin.

Next, suspend the cells in 1 ml of 20 mM Tris-HCl (pH 7.4) buffer containing 0.5 mg/ml DEAE-Dextran, 8 μg GAS-SEAP2 plasmid DNA, 140 mM NaCl, 5 mM KCl, 375 μM Na₂HPO₄.7H₂O, 1 mM MgCl₂, and 675 μM CaCl₂. Incubate at 37 degrees C. for 45 min.

Wash the cells with RPMI 1640 medium containing 10% FBS and then resuspend in 10 ml complete medium and incubate at 37 degree C. for 36 hr.

The GAS-SEAP/U937 stable cells are obtained by growing the cells in 400 μg/ml G418. The G418-free medium is used for routine growth but every one to two months, the cells should be re-grown in 400 μg/ml G418 for couple of passages.

These cells are tested by harvesting 1×10⁸ cells (this is enough for ten 96-well plates assay) and wash with PBS. Suspend the cells in 200 ml above described growth medium, with a final density of 5×10⁵ cells/ml. Plate 2000 cells per well in the 96-well plate (or 1×10⁵ cells/well).

Add 50 μl of the supernatant prepared by the protocol described in Example 30. Incubate at 37 degree C. for 48 to 72 hr. As a positive control, 100 Unit/ml interferon gamma can be used which is known to activate U937 cells. Over 30 fold induction is typically observed in the positive control wells. SEAP assay the supernatant according to the protocol described in Example 36.

Example 34 High-Throughput Screening Assay Identifying Neuronal Activity

When cells undergo differentiation and proliferation, a group of genes are activated through many different signal transduction pathways. One of these genes, EGR1 (early growth response gene 1), is induced in various tissues and cell types upon activation. The promoter of EGR1 is responsible for such induction. Using the EGR1 promoter linked to reporter molecules, activation of cells can be assessed by polypeptide of the present invention.

Particularly, the following protocol is used to assess neuronal activity in PC12 cell lines. PC12 cells (rat phenochromocytoma cells) are known to proliferate and/or differentiate by activation with a number of mitogens, such as TPA (tetradecanoyl phorbol acetate), NGF (nerve growth factor), and EGF (epidermal growth factor). The EGR1 gene expression is activated during this treatment. Thus, by stably transfecting PC12 cells with a construct containing an EGR promoter linked to SEAP reporter, activation of PC12 cells by polypeptide of the present invention can be assessed.

The EGR/SEAP reporter construct can be assembled by the following protocol. The EGR-1 promoter sequence (−633 to +1)(Sakamoto K et al., Oncogene 6:867-871 (1991)) can be PCR amplified from human genomic DNA using the following primers:

(SEQ ID NO: 6) 5′ GCGCTCGAGGGATGACAGCGATAGAACCCCGG-3′ (SEQ ID NO: 7) 5′ GCGAAGCTTCGCGACTCCCCGGATCCGCCTC-3′

Using the GAS:SEAP/Neo vector produced in Example 31, EGR1 amplified product can then be inserted into this vector. Linearize the GAS:SEAP/Neo vector using restriction enzymes XhoI/HindIII, removing the GAS/SV40 stuffer. Restrict the EGR1 amplified product with these same enzymes. Ligate the vector and the EGR1 promoter.

To prepare 96 well-plates for cell culture, two mls of a coating solution (1:30 dilution of collagen type I (Upstate Biotech Inc. Cat#08-115) in 30% ethanol (filter sterilized)) is added per one 10 cm plate or 50 ml per well of the 96-well plate, and allowed to air dry for 2 hr.

PC12 cells are routinely grown in RPMI-1640 medium (Bio Whittaker) containing 10% horse serum (JRH BIOSCIENCES, Cat. # 12449-78P), 5% heat-inactivated fetal bovine serum (FBS) supplemented with 100 units/ml penicillin and 100 μg/ml streptomycin on a precoated 10 cm tissue culture dish. One to four split is done every three to four days. Cells are removed from the plates by scraping and resuspended with pipetting up and down for more than 15 times.

Transfect the EGR/SEAP/Neo construct into PC12 using the LIPOFECTAMINE™ protocol described in Example 30. EGR-SEAP/PC12 stable cells are obtained by growing the cells in 300 μg/ml G418. The G418-free medium is used for routine growth but every one to two months, the cells should be re-grown in 300 μg/ml G418 for couple of passages.

To assay for neuronal activity, a 10 cm plate with cells around 70 to 80% confluent is screened by removing the old medium. Wash the cells once with PBS (Phosphate buffered saline). Then starve the cells in low serum medium (RPMI-1640 containing 1% horse serum and 0.5% FBS with antibiotics) overnight.

The next morning, remove the medium and wash the cells with PBS. Scrape off the cells from the plate, suspend the cells well in 2 ml low serum medium. Count the cell number and add more low serum medium to reach final cell density as 5×10⁵ cells/ml.

Add 200 μl of the cell suspension to each well of 96-well plate (equivalent to 1×10⁵ cells/well). Add 50 μl supernatant produced by Example 30, 37 degree C. for 48 to 72 hr. As a positive control, a growth factor known to activate PC12 cells through EGR can be used, such as 50 ng/μl of Neuronal Growth Factor (NGF). Over fifty-fold induction of SEAP is typically seen in the positive control wells. SEAP assay the supernatant according to Example 36.

Example 35 High-Throughput Screening Assay for T-cell Activity

NF-KB (Nuclear Factor KB) is a transcription factor activated by a wide variety of agents including the inflammatory cytokines IL-1 and TNF, CD30 and CD40, lymphotoxin-alpha and lymphotoxin-beta, by exposure to LPS or thrombin, and by expression of certain viral gene products. As a transcription factor, NF-KB regulates the expression of genes involved in immune cell activation, control of apoptosis (NF-KB appears to shield cells from apoptosis), B and T-cell development, anti-viral and antimicrobial responses, and multiple stress responses.

In non-stimulated conditions, NF-KB is retained in the cytoplasm with I-KB (Inhibitor KB). However, upon stimulation, I-KB is phosphorylated and degraded, causing NF-KB to shuttle to the nucleus, thereby activating transcription of target genes. Target genes activated by NF-KB include IL-2, IL-6, GM-CSF, ICAM-1 and class 1 MHC.

Due to its central role and ability to respond to a range of stimuli, reporter constructs utilizing the NF-KB promoter element are used to screen the supernatants produced in Example 30. Activators or inhibitors of NF-KB would be useful in detecting, preventing, diagnosing, prognosticating, treating, and/or ameliorating diseases. For example, inhibitors of NF-KB could be used to treat those diseases related to the acute or chronic activation of NF-KB, such as rheumatoid arthritis.

To construct a vector containing the NF-KB promoter element, a PCR based strategy is employed. The upstream primer contains four tandem copies of the NF-KB binding site (GGGGACTTTCCC) (SEQ ID NO: 8), 18 by of sequence complementary to the 5′ end of the SV40 early promoter sequence, and is flanked with an XhoI site:

(SEQ ID NO: 9) 5′: GCGGCCTCGAGGGGACTTTCCCGGGGACTTTCCGGGGACTTTCCGG GACTTTCCATCCTGCCATCTCAATTAG: 3′

The downstream primer is complementary to the 3′ end of the SV40 promoter and is flanked with a Hind III site:

(SEQ ID NO: 4) 5′: GCGGCAAGCTTTTTGCAAAGCCTAGGC: 3′

PCR amplification is performed using the SV40 promoter template present in the pB-gal:promoter plasmid obtained from CLONTECH™. The resulting PCR fragment is digested with XhoI and Hind III and subcloned into BLSK2-. (STRATAGENE™) Sequencing with the T7 and T3 primers confirms the insert contains the following sequence:

(SEQ ID NO: 10) 5′: CTCGAGGGGACTTTCCCGGGGACTTTCCGGGGACTTTCCGGGACTT TCCATCTGCCATCTCAATTAGTCAGCAACCATAGTCCCGCCCCTAACTCC GCCCATCCCGCCCCTAACTCCGCCCAGTTCCGCCCATTCTCCGCCCCATG GCTGACTAATTTTTTTTATTTATGCAGAGGCCGAGGCCGCCTCGGCCTCT GAGCTATTCCAGAAGTAGTGAGGAGGCTTTTTTGGAGGCCTAGGCTTTTG CAAAAAGCTT: 3′

Next, replace the SV40 minimal promoter element present in the pSEAP2-promoter plasmid (CLONTECH™) with this NF-KB/SV40 fragment using XhoI and HindIII. However, this vector does not contain a neomycin resistance gene, and therefore, is not preferred for mammalian expression systems.

In order to generate stable mammalian cell lines, the NF-KB/SV40/SEAP cassette is removed from the above NF-KB/SEAP vector using restriction enzymes SalI and NotI, and inserted into a vector containing neomycin resistance. Particularly, the NF-KB/SV40/SEAP cassette was inserted into pGFP-1 (CLONTECH™), replacing the GFP gene, after restricting pGFP-1 with SalI and NotI.

Once NF-KB/SV40/SEAP/Neo vector is created, stable Jurkat T-cells are created and maintained according to the protocol described in Example 32. Similarly, the method for assaying supernatants with these stable Jurkat T-cells is also described in Example 32. As a positive control, exogenous TNF alpha (0.1,1, 10 ng) is added to wells H9, H10, and H11, with a 5-10 fold activation typically observed.

Example 36 Assay for SEAP Activity

As a reporter molecule for the assays described in Examples 32-35, SEAP activity is assayed using the Tropix Phospho-light Kit (Cat. BP-400) according to the following general procedure. The Tropix Phospho-light Kit supplies the Dilution, Assay, and Reaction Buffers used below.

Prime a dispenser with the 2.5× Dilution Buffer and dispense 15 μl of 2.5× dilution buffer into Optiplates containing 35 μl of a supernatant. Seal the plates with a plastic sealer and incubate at 65 degree C. for 30 min. Separate the Optiplates to avoid uneven heating.

Cool the samples to room temperature for 15 minutes. Empty the dispenser and prime with the Assay Buffer. Add 50 ml Assay Buffer and incubate at room temperature 5 min. Empty the dispenser and prime with the Reaction Buffer (see the Table below). Add 50 μl Reaction Buffer and incubate at room temperature for 20 minutes. Since the intensity of the chemiluminescent signal is time dependent, and it takes about 10 minutes to read 5 plates on a luminometer, thus one should treat 5 plates at each time and start the second set 10 minutes later.

Read the relative light unit in the luminometer. Set H12 as blank, and print the results. An increase in chemiluminescence indicates reporter activity.

Reaction Buffer Formulation: # of plates Rxn buffer diluent (ml) CSPD (ml) 10 60 3 11 65 3.25 12 70 3.5 13 75 3.75 14 80 4 15 85 4.25 16 90 4.5 17 95 4.75 18 100 5 19 105 5.25 20 110 5.5 21 115 5.75 22 120 6 23 125 6.25 24 130 6.5 25 135 6.75 26 140 7 27 145 7.25 28 150 7.5 29 155 7.75 30 160 8 31 165 8.25 32 170 8.5 33 175 8.75 34 180 9 35 185 9.25 36 190 9.5 37 195 9.75 38 200 10 39 205 10.25 40 210 10.5 41 215 10.75 42 220 11 43 225 11.25 44 230 11.5 45 235 11.75 46 240 12 47 245 12.25 48 250 12.5 49 255 12.75 50 260 13

Example 37 High-Throughput Screening Assay Identifying Changes in Small Molecule Concentration and Membrane Permeability

Binding of a ligand to a receptor is known to alter intracellular levels of small molecules, such as calcium, potassium, sodium, and pH, as well as alter membrane potential. These alterations can be measured in an assay to identify supernatants which bind to receptors of a particular cell. Although the following protocol describes an assay for calcium, this protocol can easily be modified to detect changes in potassium, sodium, pH, membrane potential, or any other small molecule which is detectable by a fluorescent probe.

The following assay uses Fluorometric Imaging Plate Reader (“FLIPR”) to measure changes in fluorescent molecules (Molecular Probes) that bind small molecules. Clearly, any fluorescent molecule detecting a small molecule can be used instead of the calcium fluorescent molecule, fluo-4 (Molecular Probes, Inc.; catalog no. F-14202), used here.

For adherent cells, seed the cells at 10,000-20,000 cells/well in a Co-star black 96-well plate with clear bottom. The plate is incubated in a CO₂ incubator for 20 hours. The adherent cells are washed two times in Biotek washer with 2000 of HBSS (Hank's Balanced Salt Solution) leaving 1000 of buffer after the final wash.

A stock solution of 1 mg/ml fluo-4 is made in 10% pluronic acid DMSO. To load the cells with fluo-4, 50 μl of 12 μg/ml fluo-4 is added to each well. The plate is incubated at 37 degrees C. in a CO₂ incubator for 60 min. The plate is washed four times in the Biotek washer with HBSS leaving 1000 of buffer.

For non-adherent cells, the cells are spun down from culture media. Cells are re-suspended to 2−5×10⁶ cells/ml with HBSS in a 50-ml conical tube. 4 μl of 1 mg/ml fluo-4 solution in 10% pluronic acid DMSO is added to each ml of cell suspension. The tube is then placed in a 37 degrees C. water bath for 30-60 min. The cells are washed twice with HBSS, resuspended to 1×10⁶ cells/ml, and dispensed into a microplate, 100 μl/well. The plate is centrifuged at 1000 rpm for 5 min. The plate is then washed once in Denley Cell Wash with 200 μl, followed by an aspiration step to 1000 final volume.

For a non-cell based assay, each well contains a fluorescent molecule, such as fluo-4. The supernatant is added to the well, and a change in fluorescence is detected.

To measure the fluorescence of intracellular calcium, the FLIPR is set for the following parameters: (1) System gain is 300-800 mW; (2) Exposure time is 0.4 second; (3) Camera F/stop is F/2; (4) Excitation is 488 nm; (5) Emission is 530 nm; and (6) Sample addition is 50 μl. Increased emission at 530 nm indicates an extracellular signaling event caused by the a molecule, either polypeptide of the present invention or a molecule induced by polypeptide of the present invention, which has resulted in an increase in the intracellular Ca⁺⁺ concentration.

Example 38 High-Throughput Screening Assay Identifying Tyrosine Kinase Activity

The Protein Tyrosine Kinases (PTK) represent a diverse group of transmembrane and cytoplasmic kinases. Within the Receptor Protein Tyrosine Kinase RPTK) group are receptors for a range of mitogenic and metabolic growth factors including the PDGF, FGF, EGF, NGF, HGF and Insulin receptor subfamilies. In addition there are a large family of RPTKs for which the corresponding ligand is unknown. Ligands for RPTKs include mainly secreted small proteins, but also membrane-bound and extracellular matrix proteins.

Activation of RPTK by ligands involves ligand-mediated receptor dimerization, resulting in transphosphorylation of the receptor subunits and activation of the cytoplasmic tyrosine kinases. The cytoplasmic tyrosine kinases include receptor associated tyrosine kinases of the src-family (e.g., src, yes, lck, lyn, fyn) and non-receptor linked and cytosolic protein tyrosine kinases, such as the Jak family, members of which mediate signal transduction triggered by the cytokine superfamily of receptors (e.g., the Interleukins, Interferons, GM-CSF, and Leptin).

Because of the wide range of known factors capable of stimulating tyrosine kinase activity, identifying whether polypeptide of the present invention or a molecule induced by polypeptide of the present invention is capable of activating tyrosine kinase signal transduction pathways is of interest. Therefore, the following protocol is designed to identify such molecules capable of activating the tyrosine kinase signal transduction pathways.

Seed target cells (e.g., primary keratinocytes) at a density of approximately 25,000 cells per well in a 96 well LOPRODYNE™ Silent Screen Plates purchased from Nalge Nunc (Naperville, Ill.). The plates are sterilized with two 30 minute rinses with 100% ethanol, rinsed with water and dried overnight. Some plates are coated for 2 hr with 100 ml of cell culture grade type I collagen (50 mg/ml), gelatin (2%) or polylysine (50 mg/ml), all of which can be purchased from Sigma Chemicals (St. Louis, Mo.) or 10% MATRIGEL™ purchased from Becton Dickinson (Bedford, Mass.), or calf serum, rinsed with PBS and stored at 4 degree C. Cell growth on these plates is assayed by seeding 5,000 cells/well in growth medium and indirect quantitation of cell number through use of ALAMAR BLUE™ as described by the manufacturer Alamar Biosciences, Inc. (Sacramento, Calif.) after 48 hr. Falcon plate covers #3071 from Becton Dickinson (Bedford, Mass.) are used to cover the LOPRODYNE™ Silent Screen Plates. Falcon Microtest III cell culture plates can also be used in some proliferation experiments.

To prepare extracts, A431 cells are seeded onto the nylon membranes of LOPRODYNE™ plates (20,000/200 ml/well) and cultured overnight in complete medium. Cells are quiesced by incubation in serum-free basal medium for 24 hr. After 5-20 minutes treatment with EGF (60 ng/ml) or 500 of the supernatant produced in Example 30, the medium was removed and 100 ml of extraction buffer ((20 mM HEPES pH 7.5, 0.15 M NaCl, 1% Triton X-100, 0.1% SDS, 2 mM Na3VO4, 2 mM Na4P2O7 and a cocktail of protease inhibitors (# 1836170) obtained from Boeheringer Mannheim (Indianapolis, Ind.)) is added to each well and the plate is shaken on a rotating shaker for 5 minutes at 4° C. The plate is then placed in a vacuum transfer manifold and the extract filtered through the 0.45 mm membrane bottoms of each well using house vacuum. Extracts are collected in a 96-well catch/assay plate in the bottom of the vacuum manifold and immediately placed on ice. To obtain extracts clarified by centrifugation, the content of each well, after detergent solubilization for 5 minutes, is removed and centrifuged for 15 minutes at 4 degree C. at 16,000×g.

Test the filtered extracts for levels of tyrosine kinase activity. Although many methods of detecting tyrosine kinase activity are known, one method is described here.

Generally, the tyrosine kinase activity of a supernatant is evaluated by determining its ability to phosphorylate a tyrosine residue on a specific substrate (a biotinylated peptide). Biotinylated peptides that can be used for this purpose include PSK1 (corresponding to amino acids 6-20 of the cell division kinase cdc2-p34) and PSK2 (corresponding to amino acids 1-17 of gastrin). Both peptides are substrates for a range of tyrosine kinases and are available from Boehringer Mannheim.

The tyrosine kinase reaction is set up by adding the following components in order. First, add 100 of 5 uM Biotinylated Peptide, then 100 ATP/Mg₂₊ (5 mM ATP/50 mM MgCl₂), then 10 μl of 5× Assay Buffer (40 mM imidazole hydrochloride, pH7.3, 40 mM beta-glycerophosphate, 1 mM EGTA, 100 mM MgCl₂, 5 mM MnCl₂, 0.5 mg/ml BSA), then 5 μl of Sodium Vanadate (1 mM), and then 5 μl of water. Mix the components gently and preincubate the reaction mix at 30 degree C. for 2 min. Initial the reaction by adding 10 μl of the control enzyme or the filtered supernatant.

The tyrosine kinase assay reaction is then terminated by adding 10 μl of 120 mm EDTA and place the reactions on ice.

Tyrosine kinase activity is determined by transferring 50 μl aliquot of reaction mixture to a microtiter plate (MTP) module and incubating at 37 degree C. for 20 min. This allows the streptavidin coated 96 well plate to associate with the biotinylated peptide. Wash the MTP module with 300 ul/well of PBS four times. Next add 75 of anti-phosphotyrosine antibody conjugated to horse radish peroxidase (anti-P-Tyr-POD(0.5u/ml)) to each well and incubate at 37 degree C. for one hour. Wash the well as above.

Next add 100 l of peroxidase substrate solution (Boehringer Mannheim) and incubate at room temperature for at least 5 mins (up to 30 min) Measure the absorbance of the sample at 405 nm by using ELISA reader. The level of bound peroxidase activity is quantitated using an ELISA reader and reflects the level of tyrosine kinase activity.

Example 39 High-Throughput Screening Assay Identifying Phosphorylation Activity

As a potential alternative and/or complement to the assay of protein tyrosine kinase activity described in Example 38, an assay which detects activation (phosphorylation) of major intracellular signal transduction intermediates can also be used. For example, as described below one particular assay can detect tyrosine phosphorylation of the Erk-1 and Erk-2 kinases. However, phosphorylation of other molecules, such as Raf, JNK, p38 MAP, Map kinase kinase (MEK), MEK kinase, Src, Muscle specific kinase (MuSK), IRAK, Tec, and Janus, as well as any other phosphoserine, phosphotyrosine, or phosphothreonine molecule, can be detected by substituting these molecules for Erk-1 or Erk-2 in the following assay.

Specifically, assay plates are made by coating the wells of a 96-well ELISA plate with 0.1 ml of protein G (1 ug/ml) for 2 hr at room temp, (RT). The plates are then rinsed with PBS and blocked with 3% BSA/PBS for 1 hr at RT. The protein G plates are then treated with 2 commercial monoclonal antibodies (100 ng/well) against Erk-1 and Erk-2 (1 hr at RT) (Santa Cruz Biotechnology). (To detect other molecules, this step can easily be modified by substituting a monoclonal antibody detecting any of the above described molecules.) After 3-5 rinses with PBS, the plates are stored at 4 degree C. until use.

A431 cells are seeded at 20,000/well in a 96-well LOPRODYNE™ filterplate and cultured overnight in growth medium. The cells are then starved for 48 hr in basal medium (DMEM) and then treated with EGF (6 ng/well) or 50 μl of the supernatants obtained in Example 30 for 5-20 minutes. The cells are then solubilized and extracts filtered directly into the assay plate.

After incubation with the extract for 1 hr at RT, the wells are again rinsed. As a positive control, a commercial preparation of MAP kinase (10 ng/well) is used in place of A431 extract. Plates are then treated with a commercial polyclonal (rabbit) antibody (1 ug/ml) which specifically recognizes the phosphorylated epitope of the Erk-1 and Erk-2 kinases (1 hr at RT). This antibody is biotinylated by standard procedures. The bound polyclonal antibody is then quantitated by successive incubations with Europium-streptavidin and Europium fluorescence enhancing reagent in the Wallac DELFIA instrument (time-resolved fluorescence). An increased fluorescent signal over background indicates a phosphorylation by polypeptide of the present invention or a molecule induced by polypeptide of the present invention.

Example 40 Assay for the Stimulation of Bone Marrow CD34+ Cell Proliferation

This assay is based on the ability of human CD34+ to proliferate in the presence of hematopoietic growth factors and evaluates the ability of isolated polypeptides expressed in mammalian cells to stimulate proliferation of CD34+ cells.

It has been previously shown that most mature precursors will respond to only a single signal. More immature precursors require at least two signals to respond. Therefore, to test the effect of polypeptides on hematopoietic activity of a wide range of progenitor cells, the assay contains a given polypeptide in the presence or absence of other hematopoietic growth factors. Isolated cells are cultured for 5 days in the presence of Stem Cell Factor (SCF) in combination with tested sample. SCF alone has a very limited effect on the proliferation of bone marrow (BM) cells, acting in such conditions only as a “survival” factor. However, combined with any factor exhibiting stimulatory effect on these cells (e.g., IL-3), SCF will cause a synergistic effect. Therefore, if the tested polypeptide has a stimulatory effect on hematopoietic progenitors, such activity can be easily detected. Since normal BM cells have a low level of cycling cells, it is likely that any inhibitory effect of a given polypeptide, or agonists or antagonists thereof, might not be detected. Accordingly, assays for an inhibitory effect on progenitors is preferably tested in cells that are first subjected to in vitro stimulation with SCF+IL+3, and then contacted with the compound that is being evaluated for inhibition of such induced proliferation.

Briefly, CD34+ cells are isolated using methods known in the art. The cells are thawed and resuspended in medium (QBSF 60 serum-free medium with 1% L-glutamine (500 ml) Quality Biological, Inc., Gaithersburg, Md. Cat# 160-204-101). After several gentle centrifugation steps at 200×g, cells are allowed to rest for one hour. The cell count is adjusted to 2.5×10⁵ cells/ml. During this time, 100 μl of sterile water is added to the peripheral wells of a 96-well plate. The cytokines that can be tested with a given polypeptide in this assay is rhSCF (R&D Systems, Minneapolis, Minn., Cat# 255-SC) at 50 ng/ml alone and in combination with rhSCF and rhIL-3 (R&D Systems, Minneapolis, Minn., Cat# 203-ML) at 30 ng/ml. After one hour, 10 μl of prepared cytokines, 50 μl of the supernatants prepared in Example 30 (supernatants at 1:2 dilution=50 μl) and 20 μl of diluted cells are added to the media which is already present in the wells to allow for a final total volume of 100 μl. The plates are then placed in a 37° C./5% CO₂ incubator for five days.

Eighteen hours before the assay is harvested, 0.5 μCi/well of [3H] Thymidine is added in a 10 μl volume to each well to determine the proliferation rate. The experiment is terminated by harvesting the cells from each 96-well plate to a filtermat using the Tomtec Harvester 96. After harvesting, the filtermats are dried, trimmed and placed into OMNIFILTER™ assemblies consisting of one OMNIFILTER™ plate and one OMNIFILTER™ Tray. 60 μl MICROSCINT™ is added to each well and the plate sealed with TopSeal-A press-on sealing film A bar code 15 sticker is affixed to the first plate for counting. The sealed plates are then loaded and the level of radioactivity determined via the Packard Top Count and the printed data collected for analysis. The level of radioactivity reflects the amount of cell proliferation.

The studies described in this example test the activity of a given polypeptide to stimulate bone marrow CD34+ cell proliferation. One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof. As a nonlimiting example, potential antagonists tested in this assay would be expected to inhibit cell proliferation in the presence of cytokines and/or to increase the inhibition of cell proliferation in the presence of cytokines and a given polypeptide. In contrast, potential agonists tested in this assay would be expected to enhance cell proliferation and/or to decrease the inhibition of cell proliferation in the presence of cytokines and a given polypeptide.

The ability of a gene to stimulate the proliferation of bone marrow CD34+ cells indicates that polynucleotides and polypeptides corresponding to the gene are useful for the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of disorders affecting the immune system and hematopoiesis. Representative uses are described in the “Immune Activity” and “Infectious Disease” sections above, and elsewhere herein.

Example 41 Assay for Extracellular Matrix Enhanced Cell Response (EMECR)

The objective of the Extracellular Matrix Enhanced Cell Response (EMECR) assay is to identify gene products (e.g., isolated polypeptides) that act on the hematopoietic stem cells in the context of the extracellular matrix (ECM) induced signal.

Cells respond to the regulatory factors in the context of signal(s) received from the surrounding microenvironment. For example, fibroblasts, and endothelial and epithelial stem cells fail to replicate in the absence of signals from the ECM. Hematopoietic stem cells can undergo self-renewal in the bone marrow, but not in in vitro suspension culture. The ability of stem cells to undergo self-renewal in vitro is dependent upon their interaction with the stromal cells and the ECM protein fibronectin (fn). Adhesion of cells to fn is mediated by the α₅.β₁ and α₄.β₁ integrin receptors, which are expressed by human and mouse hematopoietic stem cells. The factor(s) which integrate with the ECM environment and are responsible for stimulating stem cell self-renewal have not yet been identified. Discovery of such factors should be of great interest in gene therapy and bone marrow transplant applications

Briefly, polystyrene, non tissue culture treated, 96-well plates are coated with fn fragment at a coating concentration of 0.2 μg/cm². Mouse bone marrow cells are plated (1,000 cells/well) in 0.2 ml of serum-free medium. Cells cultured in the presence of IL-3 (5 ng/ml)+SCF (50 ng/ml) would serve as the positive control, conditions under which little self-renewal but pronounced differentiation of the stem cells is to be expected. Gene products of the invention (e.g., including, but not limited to, polynucleotides and polypeptides of the present invention, and supernatants produced in Example 30), are tested with appropriate negative controls in the presence and absence of SCF (5.0 ng/ml), where test factor supernatants represent 10% of the total assay volume. The plated cells are then allowed to grow by incubating in a low oxygen environment (5% CO₂, 7% O₂, and 88% N₂) tissue culture incubator for 7 days. The number of proliferating cells within the wells is then quantitated by measuring thymidine incorporation into cellular DNA. Verification of the positive hits in the assay will require phenotypic characterization of the cells, which can be accomplished by scaling up of the culture system and using appropriate antibody reagents against cell surface antigens and FACScan.

One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof.

If a particular polypeptide of the present invention is found to be a stimulator of hematopoietic progenitors, polynucleotides and polypeptides corresponding to the gene encoding said polypeptide may be useful for the detection, prevention, diagnosis, prognostication, treatment, and/or amelioration of disorders affecting the immune system and hematopoiesis. Representative uses are described in the “Immune Activity” and “Infectious Disease” sections above, and elsewhere herein. The gene product may also be useful in the expansion of stem cells and committed progenitors of various blood lineages, and in the differentiation and/or proliferation of various cell types.

Additionally, the polynucleotides and/or polypeptides of the gene of interest and/or agonists and/or antagonists thereof, may also be employed to inhibit the proliferation and differentiation of hematopoietic cells and therefore may be employed to protect bone marrow stem cells from chemotherapeutic agents during chemotherapy. This antiproliferative effect may allow administration of higher doses of chemotherapeutic agents and, therefore, more effective chemotherapeutic treatment.

Moreover, polynucleotides and polypeptides corresponding to the gene of interest may also be useful for the detection, prevention, diagnosis, prognostication, treat, and/or amelioration of hematopoietic related disorders such as, for example, anemia, pancytopenia, leukopenia, thrombocytopenia or leukemia since stromal cells are important in the production of cells of hematopoietic lineages. The uses include bone marrow cell ex-vivo culture, bone marrow transplantation, bone marrow reconstitution, radiotherapy or chemotherapy of neoplasia.

Example 42 Human Dermal Fibroblast and Aortic Smooth Muscle Cell Proliferation

The polypeptide of interest is added to cultures of normal human dermal fibroblasts (NHDF) and human aortic smooth muscle cells (AoSMC) and two co-assays are performed with each sample. The first assay examines the effect of the polypeptide of interest on the proliferation of normal human dermal fibroblasts (NHDF) or aortic smooth muscle cells (AoSMC). Aberrant growth of fibroblasts or smooth muscle cells is a part of several pathological processes, including fibrosis, and restenosis. The second assay examines IL6 production by both NHDF and SMC. IL6 production is an indication of functional activation. Activated cells will have increased production of a number of cytokines and other factors, which can result in a proinflammatory or immunomodulatory outcome. Assays are run with and without co-TNFa stimulation, in order to check for costimulatory or inhibitory activity.

Briefly, on day 1,96-well black plates are set up with 1000 cells/well (NHDF) or 2000 cells/well (AoSMC) in 100 μl culture media. NHDF culture media contains: Clonetics FB basal media, 1 mg/ml hFGF, 5 mg/ml insulin, 50 mg/ml gentamycin, 2% FBS, while AoSMC culture media contains Clonetics SM basal media, 0.5 μg/ml hEGF, 5 mg/ml insulin, 1 μg/ml hFGF, 50 mg/ml gentamycin, 50 μg/ml Amphotericin B, 5% FBS. After incubation at 37° C. for at least 4-5 hours culture media is aspirated and replaced with growth arrest media. Growth arrest media for NHDF contains fibroblast basal media, 50 mg/ml gentamycin, 2% FBS, while growth arrest media for AoSMC contains SM basal media, 50 mg/ml gentamycin, 50 μg/ml Amphotericin B, 0.4% FBS. Incubate at 37° C. until day 2.

On day 2, serial dilutions and templates of the polypeptide of interest are designed such that they always include media controls and known-protein controls. For both stimulation and inhibition experiments, proteins are diluted in growth arrest media. For inhibition experiments, TNFα is added to a final concentration of 2 ng/ml (NHDF) or 5 ng/ml (AoSMC). Add 1/3 vol media containing controls or polypeptides of the present invention and incubate at 37 degrees C./5% CO₂ until day 5.

Transfer 60 μl from each well to another labeled 96-well plate, cover with a plate-sealer, and store at 4 degrees C. until Day 6 (for IL6 ELISA). To the remaining 100 μl in the cell culture plate, aseptically add ALAMAR BLUE™ in an amount equal to 10% of the culture volume (100. Return plates to incubator for 3 to 4 hours. Then measure fluorescence with excitation at 530 nm and emission at 590 nm using the CYTOFLUOR™. This yields the growth stimulation/inhibition data.

On day 5, the IL6 ELISA is performed by coating a 96 well plate with 50-100 μl/well of Anti-Human IL6 Monoclonal antibody diluted in PBS, pH 7.4, incubate ON at room temperature.

On day 6, empty the plates into the sink and blot on paper towels. Prepare Assay Buffer containing PBS with 4% BSA. Block the plates with 200 μl/well of Pierce Super Block blocking buffer in PBS for 1-2 hr and then wash plates with wash buffer (PBS, 0.05% Tween-20). Blot plates on paper towels. Then add 50 μl/well of diluted Anti-Human IL-6 Monoclonal, Biotin-labeled antibody at 0.50 mg/ml. Make dilutions of IL-6 stock in media (30, 10, 3, 1, 0.3, 0 ng/ml). Add duplicate samples to top row of plate. Cover the plates and incubate for 2 hours at RT on shaker.

Plates are washed with wash buffer and blotted on paper towels. Dilute EU-labeled Streptavidin 1:1000 in Assay buffer, and add 100 μl/well. Cover the plate and incubate 1 h at RT. Plates are again washed with wash buffer and blotted on paper towels.

Add 100 μl/well of Enhancement Solution. Shake for 5 minutes. Read the plate on the Wallac DELFIA Fluorometer. Readings from triplicate samples in each assay were tabulated and averaged.

A positive result in this assay suggests AoSMC cell proliferation and that the polypeptide of the present invention may be involved in dermal fibroblast proliferation and/or smooth muscle cell proliferation. A positive result also suggests many potential uses of polypeptides, polynucleotides, agonists and/or antagonists of the polynucleotide/polypeptide of the present invention which gives a positive result. For example, inflammation and immune responses, wound healing, and angiogenesis, as detailed throughout this specification. Particularly, polypeptides of the present invention and polynucleotides of the present invention may be used in wound healing and dermal regeneration, as well as the promotion of vasculogenesis, both of the blood vessels and lymphatics. The growth of vessels can be used in the treatment of, for example, cardiovascular diseases. Additionally, antagonists of polypeptides and polynucleotides of the invention may be useful in treating diseases, disorders, and/or conditions which involve angiogenesis by acting as an anti-vascular agent (e.g., anti-angiogenesis). These diseases, disorders, and/or conditions are known in the art and/or are described herein, such as, for example, malignancies, solid tumors, benign tumors, for example hemangiomas, acoustic neuromas, neurofibromas, trachomas, and pyogenic granulomas; artheroscleric plaques; ocular angiogenic diseases, for example, diabetic retinopathy, retinopathy of prematurity, macular degeneration, corneal graft rejection, neovascular glaucoma, retrolental fibroplasia, rubeosis, retinoblastoma, uvietis and Pterygia (abnormal blood vessel growth) of the eye; rheumatoid arthritis; psoriasis; delayed wound healing; endometriosis; vasculogenesis; granulations; hypertrophic scars (keloids); nonunion fractures; scleroderma; trachoma; vascular adhesions; myocardial angiogenesis; coronary collaterals; cerebral collaterals; arteriovenous malformations; ischemic limb angiogenesis; Osler-Webber Syndrome; plaque neovascularization; telangiectasia; hemophiliac joints; angiofibroma; fibromuscular dysplasia; wound granulation; Crohn's disease; and atherosclerosis. Moreover, antagonists of polypeptides and polynucleotides of the invention may be useful in treating anti-hyperproliferative diseases and/or anti-inflammatory known in the art and/or described herein.

One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof.

Example 43 Cellular Adhesion Molecule (CAM) Expression on Endothelial Cells

The recruitment of lymphocytes to areas of inflammation and angiogenesis involves specific receptor-ligand interactions between cell surface adhesion molecules (CAMs) on lymphocytes and the vascular endothelium. The adhesion process, in both normal and pathological settings, follows a multi-step cascade that involves intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and endothelial leukocyte adhesion molecule-1 (E-selectin) expression on endothelial cells (EC). The expression of these molecules and others on the vascular endothelium determines the efficiency with which leukocytes may adhere to the local vasculature and extravasate into the local tissue during the development of an inflammatory response. The local concentration of cytokines and growth factor participate in the modulation of the expression of these CAMs.

Briefly, endothelial cells (e.g., Human Umbilical Vein Endothelial cells (HUVECs)) are grown in a standard 96 well plate to confluence, growth medium is removed from the cells and replaced with 100 μl of 199 Medium (10% fetal bovine serum (FBS)). Samples for testing and positive or negative controls are added to the plate in triplicate (in 10 μl volumes). Plates are then incubated at 37° C. for either 5 h (selectin and integrin expression) or 24 h (integrin expression only). Plates are aspirated to remove medium and 100 μl of 0.1% paraformaldehyde-PBS (with Ca++ and Mg++) is added to each well. Plates are held at 4° C. for 30 min. Fixative is removed from the wells and wells are washed 1× with PBS(+Ca,Mg)+0.5% BSA and drained. 10 μl of diluted primary antibody is added to the test and control wells. Anti-ICAM-1-Biotin, Anti-VCAM-1-Biotin and Anti-E-selectin-Biotin are used at a concentration of 10 μg/ml (1:10 dilution of 0.1 mg/ml stock antibody). Cells are incubated at 37° C. for 30 min. in a humidified environment. Wells are washed three times with PBS (+Ca,Mg)+0.5% BSA. 20 μl of diluted ExtrAvidin-Alkaline Phosphatase (1:5,000 dilution, referred to herein as the working dilution) are added to each well and incubated at 37° C. for 30 min. Wells are washed three times with PBS (+Ca, Mg)+0.5% BSA. Dissolve 1 tablet of p-Nitrophenol Phosphate pNPP per 5 ml of glycine buffer (pH 10.4). 100 μl of pNPP substrate in glycine buffer is added to each test well. Standard wells in triplicate are prepared from the working dilution of the ExtrAvidin-Alkaline Phosphotase in glycine buffer: 1:5,000 (10⁰)>10^(−0.5)>10⁻¹>10^(−1.5).5 μl of each dilution is added to triplicate wells and the resulting AP content in each well is 5.50 ng, 1.74 ng, 0.55 ng, 0.18 ng. 100 μl of pNNP reagent is then added to each of the standard wells. The plate is incubated at 37° C. for 4 h. A volume of 50 μl of 3M NaOH is added to all wells. The plate is read on a plate reader at 405 nm using the background subtraction option on blank wells filled with glycine buffer only. Additionally, the template is set up to indicate the concentration of AP-conjugate in each standard well [5.50 ng; 1.74 ng; 0.55 ng; 0.18 ng]. Results are indicated as amount of bound AP-conjugate in each sample.

Example 44 ALAMAR BLUE™ Endothelial Cells Proliferation Assay

This assay may be used to quantitatively determine protein mediated inhibition of bFGF-induced proliferation of Bovine Lymphatic Endothelial Cells (LECs), Bovine Aortic Endothelial Cells (BAECs) or Human Microvascular Uterine Myometrial Cells (UTMECs). This assay incorporates a fluorometric growth indicator based on detection of metabolic activity. A standard ALAMAR BLUE™ Proliferation Assay is prepared in EGM-2MV with 10 ng/ml of bFGF added as a source of endothelial cell stimulation. This assay may be used with a variety of endothelial cells with slight changes in growth medium and cell concentration. Dilutions of the protein batches to be tested are diluted as appropriate. Serum-free medium (GIBCO SFM) without bFGF is used as a non-stimulated control and Angiostatin or TSP-1 are included as a known inhibitory controls.

Briefly, LEC, BAECs or UTMECs are seeded in growth media at a density of 5000 to 2000 cells/well in a 96 well plate and placed at 37 degrees C. overnight. After the overnight incubation of the cells, the growth media is removed and replaced with GIBCO EC-SFM. The cells are treated with the appropriate dilutions of the protein of interest or control protein sample(s) (prepared in SFM) in triplicate wells with additional bFGF to a concentration of 10 ng/ml. Once the cells have been treated with the samples, the plate(s) is/are placed back in the 37° C. incubator for three days. After three days 10 ml of stock ALAMAR BLUE™ (Biosource Cat# DAL1100) is added to each well and the plate(s) is/are placed back in the 37° C. incubator for four hours. The plate(s) are then read at 530 nm excitation and 590 nm emission using the CYTOFLUOR™ fluorescence reader. Direct output is recorded in relative fluorescence units.

ALAMAR BLUE™ is an oxidation-reduction indicator that both fluoresces and changes color in response to chemical reduction of growth medium resulting from cell growth. As cells grow in culture, innate metabolic activity results in a chemical reduction of the immediate surrounding environment. Reduction related to growth causes the indicator to change from oxidized (non-fluorescent blue) form to reduced (fluorescent red) form (i.e., stimulated proliferation will produce a stronger signal and inhibited proliferation will produce a weaker signal and the total signal is proportional to the total number of cells as well as their metabolic activity). The background level of activity is observed with the starvation medium alone. This is compared to the output observed from the positive control samples (bFGF in growth medium) and protein dilutions.

Example 45 Detection of Inhibition of a Mixed Lymphocyte Reaction

This assay can be used to detect and evaluate inhibition of a Mixed Lymphocyte Reaction (MLR) by gene products (e.g., isolated polypeptides) Inhibition of a MLR may be due to a direct effect on cell proliferation and viability, modulation of costimulatory molecules on interacting cells, modulation of adhesiveness between lymphocytes and accessory cells, or modulation of cytokine production by accessory cells. Multiple cells may be targeted by these polypeptides since the peripheral blood mononuclear fraction used in this assay includes T, B and natural killer lymphocytes, as well as monocytes and dendritic cells.

Polypeptides of interest found to inhibit the MLR may find application in diseases associated with lymphocyte and monocyte activation or proliferation. These include, but are not limited to, diseases such as asthma, arthritis, diabetes, inflammatory skin conditions, psoriasis, eczema, systemic lupus erythematosus, multiple sclerosis, glomerulonephritis, inflammatory bowel disease, crohn's disease, ulcerative colitis, arteriosclerosis, cirrhosis, graft vs. host disease, host vs. graft disease, hepatitis, leukemia and lymphoma.

Briefly, PBMCs from human donors are purified by density gradient centrifugation using Lymphocyte Separation Medium (LSM®, density 1.0770 g/ml, Organon Teknika Corporation, West Chester, Pa.). PBMCs from two donors are adjusted to 2×10⁶ cells/ml in RPMI-1640 (Life Technologies, Grand Island, N.Y.) supplemented with 10% FCS and 2 mM glutamine. PBMCs from a third donor is adjusted to 2×10⁵ cells/ml. Fifty microliters of PBMCs from each donor is added to wells of a 96-well round bottom microtiter plate. Dilutions of test materials (50 μl) is added in triplicate to microtiter wells. Test samples (of the protein of interest) are added for final dilution of 1:4; rhulL-2 (R&D Systems, Minneapolis, Minn., catalog number 202-IL) is added to a final concentration of 1 μg/ml; anti-CD4 mAb (R&D Systems, clone 34930.11, catalog number MAB379) is added to a final concentration of 10 μg/ml. Cells are cultured for 7-8 days at 37° C. in 5% CO₂, and 1 μC of [³H] thymidine is added to wells for the last 16 hrs of culture. Cells are harvested and thymidine incorporation determined using a Packard TopCount. Data is expressed as the mean and standard deviation of triplicate determinations.

Samples of the protein of interest are screened in separate experiments and compared to the negative control treatment, anti-CD4 mAb, which inhibits proliferation of lymphocytes and the positive control treatment, IL-2 (either as recombinant material or supernatant), which enhances proliferation of lymphocytes.

One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof.

Example 46 Assays for Protease Activity

The following assay may be used to assess protease activity of the polypeptides of the invention.

Gelatin and casein zymography are performed essentially as described (Heusen et al., Anal. Biochem., 102:196-202 (1980); Wilson et al., Journal of Urology, 149:653-658 (1993)). Samples are run on 10% polyacryamide/0.1% SDS gels containing 1% gelain orcasein, soaked in 2.5% triton at room temperature for 1 hour, and in 0.1M glycine, pH 8.3 at 37° C. 5 to 16 hours. After staining in amido black areas of proteolysis appear as clear areas against the blue-black background. Trypsin (Sigma T8642) is used as a positive control.

Protease activity is also determined by monitoring the cleavage of n-a-benzoyl-L-arginine ethyl ester (BAEE) (Sigma B-4500. Reactions are set up in (25 mMNaPO₄,1 mM EDTA, and 1 mM BAEE), pH 7.5. Samples are added and the change in absorbance at 260 nm is monitored on the Beckman DU-6 spectrophotometer in the time-drive mode. Trypsin is used as a positive control.

Additional assays based upon the release of acid-soluble peptides from casein or hemoglobin measured as absorbance at 280 nm or colorimetrically using the Folin method are performed as described in Bergmeyer, et al., Methods of Enzymatic Analysis, 5 (1984). Other assays involve the solubilization of chromogenic substrates (Ward, Applied Science, 251-317 (1983)).

Example 47 Identifying Serine Protease Substrate Specificity

Methods known in the art or described herein may be used to determine the substrate specificity of the polypeptides of the present invention having serine protease activity. A preferred method of determining substrate specificity is by the use of positional scanning synthetic combinatorial libraries as described in GB 2 324 529 (incorporated herein in its entirety).

Example 48 Ligand Binding Assays

The following assay may be used to assess ligand binding activity of the polypeptides of the invention.

Ligand binding assays provide a direct method for ascertaining receptor pharmacology and are adaptable to a high throughput format. The purified ligand for a polypeptide is radiolabeled to high specific activity (50-2000 Ci/mmol) for binding studies. A determination is then made that the process of radiolabeling does not diminish the activity of the ligand towards its polypeptide. Assay conditions for buffers, ions, pH and other modulators such as nucleotides are optimized to establish a workable signal to noise ratio for both membrane and whole cell polypeptide sources. For these assays, specific polypeptide binding is defined as total associated radioactivity minus the radioactivity measured in the presence of an excess of unlabeled competing ligand. Where possible, more than one competing ligand is used to define residual nonspecific binding.

Example 49 Functional Assay in Xenopus Oocytes

Capped RNA transcripts from linearized plasmid templates encoding the polypeptides of the invention are synthesized in vitro with RNA polymerases in accordance with standard procedures. In vitro transcripts are suspended in water at a final concentration of 0.2 mg/ml. Ovarian lobes are removed from adult female toads, Stage V defolliculated oocytes are obtained, and RNA transcripts (10 ng/oocytc) are injected in a 50 nl bolus using a microinjection apparatus. Two electrode voltage clamps are used to measure the currents from individual Xenopus oocytes in response polypeptides and polypeptide agonist exposure. Recordings are made in Ca2+ free Barth's medium at room temperature. The Xenopus system can be used to screen known ligands and tissue/cell extracts for activating ligands.

Example 50 Microphysiometric Assays

Activation of a wide variety of secondary messenger systems results in extrusion of small amounts of acid from a cell. The acid formed is largely as a result of the increased metabolic activity required to fuel the intracellular signaling process. The pH changes in the media surrounding the cell are very small but are detectable by the CYTOSENSOR microphysiometer (Molecular Devices Ltd., Menlo Park, Calif.). The CYTOSENSOR is thus capable of detecting the activation of polypeptide that is coupled to an energy utilizing intracellular signaling pathway.

Example 51 Extract/Cell Supernatant Screening

A large number of mammalian receptors exist for which there remains, as yet, no cognate activating ligand (agonist). Thus, active ligands for these receptors may not be included within the ligands banks as identified to date. Accordingly, the polypeptides of the invention can also be functionally screened (using calcium, cAMP, microphysiometer, oocyte electrophysiology, etc., functional screens) against tissue extracts to identify its natural ligands. Extracts that produce positive functional responses can be sequentially subfractionated until an activating ligand is isolated and identified.

Example 52 Calcium and cAMP Functional Assays

Seven transmembrane receptors which are expressed in HEK 293 cells have been shown to be coupled functionally to activation of PLC and calcium mobilization and/or cAMP stimulation or inhibition. Basal calcium levels in the HEK 293 cells in receptor-transfected or vector control cells were observed to be in the normal, 100 nM to 200 nM, range. HEK 293 cells expressing recombinant receptors are loaded with fura 2 and in a single day >150 selected ligands or tissue/cell extracts are evaluated for agonist induced calcium mobilization. Similarly, HEK 293 cells expressing recombinant receptors are evaluated for the stimulation or inhibition of cAMP production using standard cAMP quantitation assays. Agonists presenting a calcium transient or cAMP fluctuation are tested in vector control cells to determine if the response is unique to the transfected cells expressing receptor.

Example 53 ATP-binding Assay

The following assay may be used to assess ATP-binding activity of polypeptides of the invention.

ATP-binding activity of the polypeptides of the invention may be detected using the ATP-binding assay described in U.S. Pat. No. 5,858,719, which is herein incorporated by reference in its entirety. Briefly, ATP-binding to polypeptides of the invention is measured via photoaffinity labeling with 8-azido-ATP in a competition assay. Reaction mixtures containing 1 mg/ml of the ABC transport protein of the present invention are incubated with varying concentrations of ATP, or the non-hydrolyzable ATP analog adenyl-5′-imidodiphosphate for 10 minutes at 4° C. A mixture of 8-azido-ATP (Sigma Chem. Corp., St. Louis, Mo.) plus 8-azido-ATP (³²P-ATP) (5 mCi/μmol, ICN, Irvine Calif.) is added to a final concentration of 100 μM and 0.5 ml aliquots are placed in the wells of a porcelain spot plate on ice. The plate is irradiated using a short wave 254 nm UV lamp at a distance of 2.5 cm from the plate for two one-minute intervals with a one-minute cooling interval in between. The reaction is stopped by addition of dithiothreitol to a final concentration of 2 mM. The incubations are subjected to SDS-PAGE electrophoresis, dried, and autoradiographed. Protein bands corresponding to the particular polypeptides of the invention are excised, and the radioactivity quantified. A decrease in radioactivity with increasing ATP or adenly-5′-imidodiphosphate provides a measure of ATP affinity to the polypeptides.

Example 54 Small Molecule Screening

This invention is particularly useful for screening therapeutic compounds by using the polypeptides of the invention, or binding fragments thereof, in any of a variety of drug screening techniques. The polypeptide or fragment employed in such a test may be affixed to a solid support, expressed on a cell surface, free in solution, or located intracellularly. One method of drug screening utilizes eukaryotic or prokaryotic host cells which are stably transformed with recombinant nucleic acids expressing the polypeptide or fragment. Drugs are screened against such transformed cells in competitive binding assays. One may measure, for example, the formulation of complexes between the agent being tested and polypeptide of the invention.

Thus, the present invention provides methods of screening for drugs or any other agents which affect activities mediated by the polypeptides of the invention. These methods comprise contacting such an agent with a polypeptide of the invention or fragment thereof and assaying for the presence of a complex between the agent and the polypeptide or fragment thereof, by methods well known in the art. In such a competitive binding assay, the agents to screen are typically labeled. Following incubation, free agent is separated from that present in bound form, and the amount of free or uncomplexed label is a measure of the ability of a particular agent to bind to the polypeptides of the invention.

Another technique for drug screening provides high throughput screening for compounds having suitable binding affinity to the polypeptides of the invention, and is described in great detail in European Patent Application 84/03564, published on Sep. 13, 1984, which is herein incorporated by reference in its entirety. Briefly stated, large numbers of different small molecule test compounds are synthesized on a solid substrate, such as plastic pins or some other surface. The test compounds are reacted with polypeptides of the invention and washed. Bound polypeptides are then detected by methods well known in the art. Purified polypeptides are coated directly onto plates for use in the aforementioned drug screening techniques. In addition, non-neutralizing antibodies may be used to capture the peptide and immobilize it on the solid support.

This invention also contemplates the use of competitive drug screening assays in which neutralizing antibodies capable of binding polypeptides of the invention specifically compete with a test compound for binding to the polypeptides or fragments thereof. In this manner, the antibodies are used to detect the presence of any peptide that shares one or more antigenic epitopes with a polypeptide of the invention.

Example 55 Phosphorylation Assay

In order to assay for phosphorylation activity of the polypeptides of the invention, a phosphorylation assay as described in U.S. Pat. No. 5,958,405 (which is herein incorporated by reference) is utilized. Briefly, phosphorylation activity may be measured by phosphorylation of a protein substrate using gamma-labeled ³²P-ATP and quantitation of the incorporated radioactivity using a gamma radioisotope counter. The polypeptides of the invention are incubated with the protein substrate, ³²P-ATP, and a kinase buffer. The ³²P incorporated into the substrate is then separated from free ³²P-ATP by electrophoresis, and the incorporated ³²P is counted and compared to a negative control. Radioactivity counts above the negative control are indicative of phosphorylation activity of the polypeptides of the invention.

Example 56 Detection of Phosphorylation Activity (Activation) of the Polypeptides of the Invention in the Presence of Polypeptide Ligands

Methods known in the art or described herein may be used to determine the phosphorylation activity of the polypeptides of the invention. A preferred method of determining phosphorylation activity is by the use of the tyrosine phosphorylation assay as described in U.S. Pat. No. 5,817,471 (incorporated herein by reference).

Example 57 Identification of Signal Transduction Proteins that Interact With Polypeptides of the Present Invention

The purified polypeptides of the invention are research tools for the identification, characterization and purification of additional signal transduction pathway proteins or receptor proteins. Briefly, labeled polypeptides of the invention are useful as reagents for the purification of molecules with which it interacts. In one embodiment of affinity purification, polypeptides of the invention are covalently coupled to a chromatography column. Cell-free extract derived from putative target cells, such as carcinoma tissues, is passed over the column, and molecules with appropriate affinity bind to the polypeptides of the invention. The protein complex is recovered from the column, dissociated, and the recovered molecule subjected to N-terminal protein sequencing. This amino acid sequence is then used to identify the captured molecule or to design degenerate oligonucleotide probes for cloning the relevant gene from an appropriate cDNA library.

Example 58 IL-6 Bioassay

To test the proliferative effects of the polypeptides of the invention, the IL-6 Bioassay as described by Marz et al. is utilized (Proc. Natl. Acad. Sci., U.S.A., 95:3251-56 (1998), which is herein incorporated by reference). Briefly, IL-6 dependent B9 murine cells are washed three times in IL-6 free medium and plated at a concentration of 5,000 cells per well in 50 μl, and 50 μl of the IL-6-like polypeptide is added. After 68 hrs. at 37° C., the number of viable cells is measured by adding the tetrazolium salt thiazolyl blue (MTT) and incubating for a further 4 hrs. at 37° C. B9 cells are lysed by SDS and optical density is measured at 570 nm. Controls containing IL-6 (positive) and no cytokine (negative) are utilized. Enhanced proliferation in the test sample(s) relative to the negative control is indicative of proliferative effects mediated by polypeptides of the invention.

Example 59 Support of Chicken Embryo Neuron Survival

To test whether sympathetic neuronal cell viability is supported by polypeptides of the invention, the chicken embryo neuronal survival assay of Senaldi et al is utilized (Proc. Natl. Acad. Sci., U.S.A., 96:11458-63 (1998), which is herein incorporated by reference). Briefly, motor and sympathetic neurons are isolated from chicken embryos, resuspended in L15 medium (with 10% FCS, glucose, sodium selenite, progesterone, conalbumin, putrescine, and insulin; Life Technologies, Rockville, Md.) and Dulbecco's modified Eagles medium [with 10% FCS, glutamine, penicillin, and 25 mM Hepes buffer (pH 7.2); Life Technologies, Rockville, Md.], respectively, and incubated at 37° C. in 5% CO₂ in the presence of different concentrations of the purified IL-6-like polypeptide, as well as a negative control lacking any cytokine. After 3 days, neuron survival is determined by evaluation of cellular morphology, and through the use of the colorimetric assay of Mosmann (Mosmann, T., J. Immunol. Methods, 65:55-63 (1983)). Enhanced neuronal cell viability as compared to the controls lacking cytokine is indicative of the ability of the inventive purified IL-6-like polypeptide(s) to enhance the survival of neuronal cells.

Example 60 Assay for Phosphatase Activity

The following assay may be used to assess serine/threonine phosphatase (PTPase) activity of the polypeptides of the invention.

In order to assay for serine/threonine phosphatase (PTPase) activity, assays can be utilized which are widely known to those skilled in the art. For example, the serine/threonine phosphatase (PSPase) activity is measured using a PSPase assay kit from New England Biolabs, Inc. Myelin basic protein (MyBP), a substrate for PSPase, is phosphorylated on serine and threonine residues with cAMP-dependent Protein Kinase in the presence of [³²P]ATP. Protein serine/threonine phosphatase activity is then determined by measuring the release of inorganic phosphate from ³²P-labeled MyBP.

Example 61 Interaction of Serine/Threonine Phosphatases with other Proteins

The polypeptides of the invention with serine/threonine phosphatase activity as determined in Example 60 are research tools for the identification, characterization and purification of additional interacting proteins or receptor proteins, or other signal transduction pathway proteins. Briefly, labeled polypeptide(s) of the invention is useful as a reagent for the purification of molecules with which it interacts. In one embodiment of affinity purification, polypeptide of the invention is covalently coupled to a chromatography column. Cell-free extract derived from putative target cells, such as neural or liver cells, is passed over the column, and molecules with appropriate affinity bind to the polypeptides of the invention. The polypeptides of the invention-complex is recovered from the column, dissociated, and the recovered molecule subjected to N-terminal protein sequencing. This amino acid sequence is then used to identify the captured molecule or to design degenerate oligonucleotide probes for cloning the relevant gene from an appropriate cDNA library.

Example 62 Assaying for Heparanase Activity

In order to assay for heparanase activity of the polypeptides of the invention, the heparanase assay described by Vlodaysky et al is utilized (Vlodaysky, I., et al., Nat. Med., 5:793-802 (1999)). Briefly, cell lysates, conditioned media or intact cells (1×10⁶ cells per 35-mm dish) are incubated for 18 hrs at 37° C., pH 6.2-6.6, with ³⁵S-labeled ECM or soluble ECM derived peak I proteoglycans. The incubation medium is centrifuged and the supernatant is analyzed by gel filtration on a Sepharose CL-6B column (0.9×30 cm). Fractions are eluted with PBS and their radioactivity is measured. Degradation fragments of heparan sulfate side chains are eluted from Sepharose 6B at 0.5<K_(av)<0.8 (peak II). Each experiment is done at least three times. Degradation fragments corresponding to “peak II,” as described by Vlodaysky et al., is indicative of the activity of the polypeptides of the invention in cleaving heparan sulfate.

Example 63 Immobilization of Biomolecules

This example provides a method for the stabilization of polypeptides of the invention in non-host cell lipid bilayer constucts (see, e.g., Bieri et al., Nature Biotech 17:1105-1108 (1999), hereby incorporated by reference in its entirety herein) that can be adapted for the study of polypeptides of the invention in the various functional assays described above. Briefly, carbohydrate-specific chemistry for biotinylation is used to confine a biotin tag to the extracellular domain of the polypeptides of the invention, thus allowing uniform orientation upon immobilization. A 50 uM solution of polypeptides of the invention in washed membranes is incubated with 20 mM NaIO4 and 1.5 mg/ml (4 mM) BACH or 2 mg/ml (7.5 mM) biotin-hydrazide for 1 hr at room temperature (reaction volume, 150u1). Then the sample is dialyzed (Pierce Slidealizer Cassett, 10 kDa cutoff; Pierce Chemical Co., Rockford Ill.) at 4° C. first for 5 h, exchanging the buffer after each hour, and finally for 12 h against 500 ml buffer R (0.15 M NaCl, 1 mM MgCl₂, 10 mM sodium phosphate, pH7). Just before addition into a cuvette, the sample is diluted 1:5 in buffer ROG50 (Buffer R supplemented with 50 mM octylglucoside).

Example 64 TAQMAN

Quantitative PCR (QPCR). Total RNA from cells in culture are extracted by Trizol separation as recommended by the supplier (LifeTechnologies). (Total RNA is treated with DNase I (Life Technologies) to remove any contaminating genomic DNA before reverse transcription.) Total RNA (50 ng) is used in a one-step, 50 ul, RT-QPCR, consisting of Taqman Buffer A (Perkin-Elmer; 50 mM KC1/10 mM Tris, pH 8.3), 5.5 mM MgCl₂, 240 μM each dNTP, 0.4 units RNase inhibitor (PROMEGA™), 8% glycerol, 0.012% Tween-20, 0.05% gelatin, 0.3 μM primers, 0.1 μM probe, 0.025 units Amplitaq Gold (Perkin-Elmer) and 2.5 units Superscript II reverse transcriptase (Life Technologies). As a control for genomic contamination, parallel reactions are setup without reverse transcriptase. The relative abundance of (unknown) and 18S RNAs are assessed by using the Applied Biosystems Prism 7700 Sequence Detection System (Livak, K. J., Flood, S. J., Marmaro, J., Giusti, W. & Deetz, K. (1995) PCR Methods Appl. 4, 357-362). Reactions are carried out at 48° C. for 30 min, 95° C. for 10 min, followed by 40 cycles of 95° C. for 15s, 60° C. for 1 min. Reactions are performed in triplicate.

Primers (f & r) and FRET probes sets are designed using Primer Express Software (Perkin-Elmer). Probes are labeled at the 5′-end with the reporter dye 6-FAM and on the 3′-end with the quencher dye TAMRA (Biosource International, Camarillo, Calif. or Perkin-Elmer).

Example 65 Assays for Metalloproteinase Activity

Metalloproteinases (EC 3.4.24.-) are peptide hydrolases which use metal ions, such as Zn²⁺, as the catalytic mechanism. Metalloproteinase activity of polypeptides of the present invention can be assayed according to the following methods.

Proteolysis of alpha-2-macroglobulin

To confirm protease activity, purified polypeptides of the invention are mixed with the substrate alpha-2-macroglobulin (0.2 unit/ml; Boehringer Mannheim, Germany) in 1× assay buffer (50 mM HEPES, pH 7.5, 0.2 M NaCl, 10 mM CaCl₂, 25 μM ZnCl₂ and 0.05% Brij-35) and incubated at 37° C. for 1-5 days. Trypsin is used as positive control. Negative controls contain only alpha-2-macroglobulin in assay buffer. The samples are collected and boiled in SDS-PAGE sample buffer containing 5% 2-mercaptoethanol for 5-min, then loaded onto 8% SDS-polyacrylamide gel. After electrophoresis the proteins are visualized by silver staining. Proteolysis is evident by the appearance of lower molecular weight bands as compared to the negative control.

Inhibition of alpha-2-macroglobulin proteolysis by inhibitors of metalloproteinases

Known metalloproteinase inhibitors (metal chelators (EDTA, EGTA, AND HgCl₂), peptide metalloproteinase inhibitors (TIMP-1 and TIMP-2), and commercial small molecule MMP inhibitors) are used to characterize the proteolytic activity of polypeptides of the invention. The three synthetic MMP inhibitors used are: MMP inhibitor I, [IC₅₀=1.0 μM against MMP-1 and MMP-8; IC₅₀=30 μM against MMP-9; IC₅₀=150 μM against MMP-3]; MMP-3 (stromelysin-1) inhibitor I [IC₅₀=5 μM against MMP-3], and MMP-3 inhibitor II [K_(i)=130 nM against MMP-3]; inhibitors available through Calbiochem, catalog #444250, 444218, and 444225, respectively). Briefly, different concentrations of the small molecule MMP inhibitors are mixed with purified polypeptides of the invention (50 μg/ml) in 22.9 μl of 1×HEPES buffer (50 mM HEPES, pH 7.5, 0.2 M NaCl, 10 mM CaCl₂, 25 μM ZnCl₂ and 0.05% Brij-35) and incubated at room temperature (24° C.) for then 7.1 μl of substrate alpha-2-macroglobulin (0.2 unit/ml) is added and incubated at 37° C. for 20-hr. The reactions are stopped by adding 4× sample buffer and boiled immediately for 5 minutes. After SDS-PAGE, the protein bands are visualized by silver stain.

Synthetic Fluorogenic Peptide Substrates Cleavage Assay

The substrate specificity for polypeptides of the invention with demonstrated metalloproteinase activity can be determined using synthetic fluorogenic peptide substrates (purchased from BACHEM Bioscience Inc). Test substrates include, M-1985, M-2225, M-2105, M-2110, and M-2255. The first four are MMP substrates and the last one is a substrate of tumor necrosis factor-α (TNF-α) converting enzyme (TACE). All the substrates are prepared in 1:1 dimethyl sulfoxide (DMSO) and water. The stock solutions are 50-500 μM. Fluorescent assays are performed by using a Perkin Elmer LS 50B luminescence spectrometer equipped with a constant temperature water bath. The excitation 2 is 328 nm and the emission 2 is 393 nm. Briefly, the assay is carried out by incubating 176 μl 1×HEPES buffer (0.2 M NaCl, 10 mM CaCl₂, 0.05% Brij-35 and 50 mM HEPES, pH 7.5) with 4 μl of substrate solution (50 μM) at 25° C. for 15 minutes, and then adding 20 μl of a purified polypeptide of the invention into the assay cuvett. The final concentration of substrate is 1 μM. Initial hydrolysis rates are monitored for 30-min.

Example 66 Characterization of the cDNA Contained in a Deposited Plasmid

The size of the cDNA insert contained in a deposited plasmid may be routinely determined using techniques known in the art, such as PCR amplification using synthetic primers hybridizable to the 3′ and 5′ ends of the cDNA sequence. For example, two primers of 17-30 nucleotides derived from each end of the cDNA (i.e., hybridizable to the absolute 5′ nucleotide or the 3′ nucleotide end of the sequence of SEQ ID NO:X, respectively) are synthesized and used to amplify the cDNA using the deposited cDNA plasmid as a template. The polymerase chain reaction is carried out under routine conditions, for instance, in 25 μl of reaction mixture with 0.5 μg of the above cDNA template. A convenient reaction mixture is 1.5-5 mM MgCl₂, 0.01% (w/v) gelatin, 20 μM each of dATP, dCTP, dGTP, dTTP, 25 μmol of each primer and 0.25 Unit of Taq polymerase. Thirty five cycles of PCR (denaturation at 94 degree C. for 1 min; annealing at 55 degree C. for 1 min; elongation at 72 degree C. for 1 min) are performed with a Perkin-Elmer Cetus automated thermal cycler. The amplified product is analyzed by agarose gel electrophoresis. The PCR product is verified to be the selected sequence by subcloning and sequencing the DNA product. It will be clear that the invention may be practiced otherwise than as particularly described in the foregoing description and examples. Numerous modifications and variations of the present invention are possible in light of the above teachings and, therefore, are within the scope of the appended claims.

Incorporation by Reference

The entire disclosure of each document cited (including patents, patent applications, journal articles, abstracts, laboratory manuals, books, or other disclosures) in the Background of the Invention, Detailed Description, and Examples is hereby incorporated herein by reference. In addition, the sequence listing submitted herewith is incorporated herein by reference in its entirety. The specification and sequence listing of each of the following U.S. and PCT applications are herein incorporated by reference in their entirety: U.S. Appln. No. 60/040,162 filed on 7 Mar. 1997, U.S. Appln. No. 60/043,576 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,601 filed on 23 May 1997, U.S. Appln. No. 60/056,845 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,580 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,599 filed on 23 May 1997, U.S. Appln. No. 60/056,664 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,314 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,632 filed on 23 May 1997, U.S. Appln. No. 60/056,892 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,568 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,595 filed on 23 May 1997, U.S. Appln. No. 60/056,632 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,578 filed on 11 Apr. 1997, U.S. Appln. No. 60/040,333 filed on 7 Mar. 1997, U.S. Appln. No. 60/043,670 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,596 filed on 23 May 1997, U.S. Appln. No. 60/056,864 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,674 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,612 filed on 23 May 1997, U.S. Appln. No. 60/056,631 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,569 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,588 filed on 23 May 1997, U.S. Appln. No. 60/056,876 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,671 filed on 11 Apr. 1997, U.S. Appln. No. 60/043,311 filed on 11 Apr. 1997, U.S. Appln. No. 60/038,621 filed on 7 Mar. 1997, U.S. Appln. No. 60/043,672 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,613 filed on 23 May 1997, U.S. Appln. No. 60/056,636 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,669 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,582 filed on 23 May 1997, U.S. Appln. No. 60/056,910 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,315 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,598 filed on 23 May 1997, U.S. Appln. No. 60/056,874 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,312 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,585 filed on 23 May 1997, U.S. Appln. No. 60/056,881 filed on 22 Aug. 1997, U.S. Appln. No. 60/043,313 filed on 11 Apr. 1997, U.S. Appln. No. 60/047,586 filed on 23 May 1997, U.S. Appln. No. 60/056,909 filed on 22 Aug. 1997, U.S. Appln. No. 60/040,161 filed on 7 Mar. 1997, U.S. Appln. No. 60/047,587 filed on 23 May 1997, U.S. Appln. No. 60/056,879 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,500 filed on 23 May 1997, U.S. Appln. No. 60/056,880 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,584 filed on 23 May 1997, U.S. Appln. No. 60/056,894 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,492 filed on 23 May 1997, U.S. Appln. No. 60/056,911 filed on 22 Aug. 1997, U.S. Appln. No. 60/040,626 filed on 7 Mar. 1997, U.S. Appln. No. 60/047,503 filed on 23 May 1997, U.S. Appln. No. 60/056,903 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,501 filed on 23 May 1997, U.S. Appln. No. 60/056,637 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,590 filed on 23 May 1997, U.S. Appln. No. 60/056,875 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,581 filed on 23 May 1997, U.S. Appln. No. 60/056,882 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,592 filed on 23 May 1997, U.S. Appln. No. 60/056,888 filed on 22 Aug. 1997, U.S. Appln. No. 60/040,334 filed on 7 Mar. 1997, U.S. Appln. No. 60/047,618 filed on 23 May 1997, U.S. Appln. No. 60/056,872 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,617 filed on 23 May 1997, U.S. Appln. No. 60/056,662 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,589 filed on 23 May 1997, U.S. Appln. No. 60/056,862 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,594 filed on 23 May 1997, U.S. Appln. No. 60/056,884 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,583 filed on 23 May 1997, U.S. Appln. No. 60/056,878 filed on 22 Aug. 1997, U.S. Appln. No. 60/040,336 filed on 7 Mar. 1997, U.S. Appln. No. 60/047,502 filed on 23 May 1997, U.S. Appln. No. 60/056,893 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,633 filed on 23 May 1997, U.S. Appln. No. 60/056,630 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,593 filed on 23 May 1997, U.S. Appln. No. 60/056,887 filed on 22 Aug. 1997, U.S. Appln. No. 60/040,163 filed on 7 Mar. 1997, U.S. Appln. No. 60/047,597 filed on 23 May 1997, U.S. Appln. No. 60/056,889 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,615 filed on 23 May 1997, U.S. Appln. No. 60/056,877 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,600 filed on 23 May 1997, U.S. Appln. No. 60/056,886 filed on 22 Aug. 1997, U.S. Appln. No. 60/047,614 filed on 23 May 1997, U.S. Appln. No. 60/056,908 filed on 22 Aug. 1997, U.S. Appln. No. 60/040,710 filed on 14 Mar. 1997, U.S. Appln. No. 60/050,934 filed on 30 May 1997, U.S. Appln. No. 60/048,100 filed on 30 May 1997, U.S. Appln. No. 60/040,762 filed on 14 Mar. 1997, U.S. Appln. No. 60/048,357 filed on 30 May 1997, U.S. Appln. No. 60/048,189 filed on 30 May 1997, U.S. Appln. No. 60/041,277 filed on 21 Mar. 1997, U.S. Appln. No. 60/048,188 filed on 30 May 1997, U.S. Appln. No. 60/048,094 filed on 30 May 1997, U.S. Appln. No. 60/048,350 filed on 30 May 1997, U.S. Appln. No. 60/048,135 filed on 30 May 1997, U.S. Appln. No. 60/042,344 filed on 21 Mar. 1997, U.S. Appln. No. 60/048,187 filed on 30 May 1997, U.S. Appln. No. 60/048,099 filed on 30 May 1997, U.S. Appln. No. 60/050,937 filed on 30 May 1997, U.S. Appln. No. 60/048,352 filed on 30 May 1997, U.S. Appln. No. 60/041,276 filed on 21 Mar. 1997, U.S. Appln. No. 60/048,069 filed on 30 May 1997, U.S. Appln. No. 60/048,131 filed on 30 May 1997, U.S. Appln. No. 60/048,186 filed on 30 May 1997, U.S. Appln. No. 60/048,095 filed on 30 May 1997, U.S. Appln. No. 60/041,281 filed on 21 Mar. 1997, U.S. Appln. No. 60/048,355 filed on 30 May 1997, U.S. Appln. No. 60/048,096 filed on 30 May 1997, U.S. Appln. No. 60/048,351 filed on 30 May 1997, U.S. Appln. No. 60/048,154 filed on 30 May 1997, U.S. Appln. No. 60/048,160 filed on 30 May 1997, U.S. Appln. No. 60/042,825 filed on 8 Apr. 1997, U.S. Appln. No. 60/048,070 filed on 30 May 1997, U.S. Appln. No. 60/042,727 filed on 8 Apr. 1997, U.S. Appln. No. 60/048,068 filed on 30 May 1997, U.S. Appln. No. 60/042,726 filed on 8 Apr. 1997, U.S. Appln. No. 60/048,184 filed on 30 May 1997, U.S. Appln. No. 60/042,728 filed on 8 Apr. 1997, U.S. Appln. No. 60/042,754 filed on 8 Apr. 1997, U.S. Appln. No. 60/048,190 filed on 30 May 1997, U.S. Appln. No. 60/044,039 filed on 30 May 1997, U.S. Appln. No. 60/048,093 filed on 30 May 1997, U.S. Appln. No. 60/048,885 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,645 filed on 5 Sep. 1997, U.S. Appln. No. 60/049,375 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,642 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,881 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,668 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,880 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,635 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,896 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,627 filed on 5 Sep. 1997, U.S. Appln. No. 60/049,020 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,667 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,876 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,666 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,895 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,764 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,884 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,643 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,894 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,769 filed on 5 Sep. 1997, U.S. Appln. No. 60/048,971 filed on 6 Jun. 1997, U.S. Appln. No. 60/057,763 filed on 5 Sep. 1997, U.S. Appln. 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LENGTHY TABLES The patent application contains a lengthy table section. A copy of the table is available in electronic form from the USPTO web site (http://seqdata.uspto.gov/?pageRequest=docDetail&DocID=US20110105726A1). An electronic copy of the table will also be available from the USPTO upon request and payment of the fee set forth in 37 CFR 1.19(b)(3). 

1-12. (canceled)
 13. An isolated antibody or fragment thereof that binds specifically to an isolated HSDEK49 polypeptide selected from the group consisting of: (a) a polypeptide consisting of amino acid residues 1 to 399 of SEQ ID NO:1373; (b) a polypeptide consisting of amino acid residues 2 to 399 of SEQ ID NO:1373; (c) a polypeptide consisting of amino acid residues 20 to 399 of SEQ ID NO:1373; (d) a polypeptide consisting of the amino acid sequence of the full-length polypeptide, which amino acid sequence is encoded by the cDNA contained in ATCC™ Deposit No:209603, which was deposited on Jan. 29, 1998; (e) a polypeptide consisting of the amino acid sequence of the HSDEK49 polypeptide, excluding the N-terminal methionine residue, which amino acid sequence is encoded by a polynucleotide contained within the cDNA contained in ATCC™ Deposit No:209603, which was deposited on Jan. 29, 1998; (f) a polypeptide consisting of the amino acid sequence of the HSDEK49 polypeptide, excluding the leader sequence of the polypeptide, which amino acid sequence is encoded by a polynucleotide contained within the cDNA contained in ATCC™ Deposit No:209603, which was deposited on Jan. 29, 1998; (g) an extracellular domain of the polypeptide consisting of amino acid residues 1 to 399 of SEQ ID NO:1373; and (h) an extracellular domain of the polypeptide, which amino acid sequence is encoded by a polynucleotide, which is contained in the cDNA contained in ATCC™ Deposit No:209603, which was deposited on Jan. 29,
 1998. 14-20. (canceled)
 21. The antibody or fragment thereof of claim 13 that specifically binds (a).
 22. The antibody or fragment thereof of claim 13 that specifically binds (b).
 23. The antibody or fragment thereof of claim 13 that specifically binds (c).
 24. The antibody or fragment thereof of claim 13 that specifically binds (d).
 25. The antibody or fragment thereof of claim 13 that specifically binds (e).
 26. The antibody or fragment thereof of claim 13 that specifically binds (f).
 27. The antibody or fragment thereof of claim 13 that specifically binds (g).
 28. The antibody or fragment thereof of claim 13 that specifically binds (h).
 29. The antibody or fragment thereof of claim 13 wherein said antibody or fragment thereof is polyclonal.
 30. The antibody or fragment thereof of claim 13 wherein said antibody or fragment thereof is monoclonal.
 31. The antibody or fragment thereof of claim 13 which is selected from the group consisting of: (a) a chimeric antibody or fragment thereof; (b) a humanized antibody or fragment thereof; (c) a single chain antibody; and (d) a Fab fragment.
 32. A hybridoma that produces the antibody or fragment thereof of claim
 13. 33. A recombinant host cell that expresses the isolated antibody or fragment thereof of claim
 13. 34. A method of making an isolated antibody or fragment thereof comprising: (a) culturing the recombinant host cell of claim 33 under conditions such that said antibody or fragment thereof is expressed; and (b) recovering said antibody or fragment thereof.
 35. An isolated antibody or fragment thereof that specifically binds a HSDEK49 polypeptide purified from a cell culture wherein said HSDEK49 polypeptide consists of amino acid residues 1 to 399 of SEQ ID NO:1373.
 36. The antibody or fragment thereof of claim 35 which is selected from the group consisting of: (a) a chimeric antibody or fragment thereof; (b) a humanized antibody or fragment thereof; (c) a single chain antibody; and (d) a Fab fragment. 